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Sample records for ataxia-telangiectasia cells exposed

  1. Chromosome aberrations in ataxia telangiectasia cells exposed to heavy ions

    Science.gov (United States)

    Kawata, T.; Cucinotta, F.; George, K.; Wu, H.; Shigematsu, N.; Furusawa, Y.; Uno, T.; Isobe, K.; Ito, H.

    Understanding of biological effects of heavy ions is important to assess healt h risk in space. One of the most important issues may be to take into account individual susceptibility. Ataxia telangiectasia (A-T) cells are known to exhibit abnormal responses to radiations but the mechanism of hyper radiosensitivity of A-T still remains unknown. We report chromosome aberrations in normal human fibroblasts and AT fibroblasts exposed to low- and high-LET radiations. A chemical-induced premature chromosome condensation (PCC) technique combined with chromosome- painting technique was applied to score chromosome aberrations in G2/M-phase cells. Following gamma irradiation, GM02052 cells were approximately 5 times more sensitive to g-rays than AG1522 cells. GM02052 cells had a much higher frequency of deletions and misrejoining than AG1522 cells. When the frequency of complex type aberrations was compared, GM02052 cells showed more than 10 times higher frequency than AG1522 cells. The results will be compared with those obtained from high-LET irradiations.

  2. Ataxia - telangiectasia

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/001394.htm Ataxia - telangiectasia To use the sharing features on this page, please enable JavaScript. Ataxia-telangiectasia is a rare childhood disease. It affects ...

  3. Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis

    OpenAIRE

    Zhe-Wei Zhang; Jing Xiao; Wei Luo; Bo-Han Wang; Ji-Min Chen

    2015-01-01

    Background: Caffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM) activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR). Methods: Immunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine...

  4. Ataxia Telangiectasia

    Science.gov (United States)

    Ataxia-telangiectasia (A-T) is a rare, inherited disease. It affects the nervous system, immune system, and ... young children, usually before age 5. They include Ataxia - trouble coordinating movements Poor balance Slurred speech Tiny, ...

  5. Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young June; Ahn, Kwang Sung; Kim, Minjeong; Kim, Min Ju; Park, Sang-Min; Ryu, Junghyun; Ahn, Jin Seop; Heo, Soon Young; Kang, Jee Hyun; Choi, You Jung [Department of Nanobiomedical Science and BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan (Korea, Republic of); Choi, Seong-Jun [Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of); Shim, Hosup, E-mail: shim@dku.edu [Department of Nanobiomedical Science and BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan (Korea, Republic of); Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of); Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of)

    2014-10-03

    Highlights: • ATM gene-targeted pigs were produced by somatic cell nuclear transfer. • A novel large animal model for ataxia telangiectasia was developed. • The new model may provide an alternative to the mouse model. - Abstract: Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human. In an attempt to provide a large animal model for A-T, we produced gene-targeted pigs with mutations in the ATM gene by somatic cell nuclear transfer. The disrupted allele in the ATM gene of cloned piglets was confirmed via PCR and Southern blot analysis. The ATM gene-targeted pigs generated in the present study may provide an alternative to the current mouse model for the study of mechanisms underlying A-T disorder and for the development of new therapies.

  6. Radiosensitive melanoma cell line with mutation of the gene for ataxia telangiectasia.

    OpenAIRE

    Ramsay, J.; Birrell, G.; K. Baumann; Bodero, A.; Parsons, P.; Lavin, M

    1998-01-01

    The human melanoma cell lines MM96L, A2058 and HT144 were examined for sensitivity to ionizing radiation and UVB radiation. HT144 demonstrated a significant increase in sensitivity to ionizing and UVB radiation compared with the MM96L and A2058 cells. Sensitivity to both agents was associated with susceptibility to apoptosis. Using a protein truncation assay, a mutation for the gene for ataxia telangiectasia (ATM) was identified in HT144 cells. This was confirmed to be a homozygous mutation b...

  7. Analysis of Residual DSBs in Ataxia-Telangiectasia Lymphoblast Cells Initiating Apoptosis

    Directory of Open Access Journals (Sweden)

    Teresa Anglada

    2016-01-01

    Full Text Available In order to examine the relationship between accumulation of residual DNA double-strand breaks (DSBs and cell death, we have used a control and an ATM (Ataxia-Telangiectasia Mutated defective cell line, as Ataxia-Telangiectasia (AT cells tend to accumulate residual DSBs at long times after damage infliction. After irradiation, AT cells showed checkpoint impairment and a fraction of cells displayed an abnormal centrosome number and tetraploid DNA content, and this fraction increased along with apoptosis rates. At all times analyzed, AT cells displayed a significantly higher rate of radiation-induced apoptosis than normal cells. Besides apoptosis, 70–85% of the AT viable cells (TUNEL-negative carried ≥10 γH2AX foci/cell, while only 12–27% of normal cells did. The fraction of AT and normal cells undergoing early and late apoptosis were isolated by flow cytometry and residual DSBs were concretely scored in these populations. Half of the γH2AX-positive AT cells undergoing early apoptosis carried ≥10 γH2AX foci/cell and this fraction increased to 75% in late apoptosis. The results suggest that retention of DNA damage-induced γH2AX foci is an indicative of lethal DNA damage, as cells undergoing apoptosis are those accumulating more DSBs. Scoring of residual γH2AX foci might function as a predictive tool to assess radiation-induced apoptosis.

  8. Cell and Molecular Biology of Ataxia Telangiectasia Heterozygous Human Mammary Epithelial Cells Irradiated in Culture

    Science.gov (United States)

    Richmond, Robert C.

    2001-01-01

    Autologous isolates of cell types from obligate heterozygotes with the autosomal disorder ataxia-telangiectasia (A-T)were used to begin a tissue culture model for assessing pathways of radiation-induced cancer formation in this target tissue. This was done by establishing cultures of stromal fibroblasts and long-term growth human mammary epithelial cells (HMEC) in standard 2-dimensional tissue culture in order to establish expression of markers detailing early steps of carcinogenesis. The presumptive breast cancer susceptibility of A-T heterozygotes as a sequel to damage caused by ionizing radiation provided reason to study expression of markers in irradiated HMEC. Findings from our study with HMEC have included determination of differences in specific protein expression amongst growth phase (e.g., log vs stationary) and growth progression (e.g., pass 7 vs pass 9), as well as differences in morphologic markers within populations of irradiated HMEC (e.g., development of multinucleated cells).

  9. Small-molecule inhibitors of Ataxia Telangiectasia and Rad3 related kinase (ATR) sensitize lymphoma cells to UVA radiation

    DEFF Research Database (Denmark)

    Biskup, Edyta; Naym, David Gram; Gniadecki, Robert

    2016-01-01

    and require more aggressive therapies. OBJECTIVE: The aim of this project was to investigate whether inhibition of Ataxia Telangiectasia and Rad3 related kinase (ATR) may enhance efficacy of phototherapy. METHODS: CTCL cell lines (MyLa2000, SeAx and Mac2a) served as in vitro cell models. ATR and Chk1 were...

  10. Control of cell respiration by nitric oxide in Ataxia Telangiectasia lymphoblastoid cells.

    Science.gov (United States)

    Masci, Alessandra; Mastronicola, Daniela; Arese, Marzia; Piane, Maria; De Amicis, Andrea; Blanck, Thomas J J; Chessa, Luciana; Sarti, Paolo

    2008-01-01

    Ataxia Telangiectasia (AT) patients are particularly sensitive to oxidative-nitrosative stress. Nitric oxide (NO) controls mitochondrial respiration via the reversible inhibition of complex IV. The mitochondrial response to NO of AT lymphoblastoid cells was investigated. Cells isolated from three patients and three intrafamilial healthy controls were selected showing within each group a normal diploid karyotype and homogeneous telomere length. Different complex IV NO-inhibition patterns were induced by varying the electron flux through the respiratory chain, using exogenous cell membrane permeable electron donors. Under conditions of high electron flux the mitochondrial NO inhibition of respiration was greater in AT than in control cells (P< or =0.05). This property appears peculiar to AT, and correlates well to the higher concentration of cytochrome c detected in the AT cells. This finding is discussed on the basis of the proposed mechanism of reaction of NO with complex IV. It is suggested that the peculiar response of AT mitochondria to NO stress may be relevant to the mitochondrial metabolism of AT patients.

  11. Donor-derived brain tumor following neural stem cell transplantation in an ataxia telangiectasia patient.

    Directory of Open Access Journals (Sweden)

    Ninette Amariglio

    2009-02-01

    Full Text Available BACKGROUND: Neural stem cells are currently being investigated as potential therapies for neurodegenerative diseases, stroke, and trauma. However, concerns have been raised over the safety of this experimental therapeutic approach, including, for example, whether there is the potential for tumors to develop from transplanted stem cells. METHODS AND FINDINGS: A boy with ataxia telangiectasia (AT was treated with intracerebellar and intrathecal injection of human fetal neural stem cells. Four years after the first treatment he was diagnosed with a multifocal brain tumor. The biopsied tumor was diagnosed as a glioneuronal neoplasm. We compared the tumor cells and the patient's peripheral blood cells by fluorescent in situ hybridization using X and Y chromosome probes, by PCR for the amelogenin gene X- and Y-specific alleles, by MassArray for the ATM patient specific mutation and for several SNPs, by PCR for polymorphic microsatellites, and by human leukocyte antigen (HLA typing. Molecular and cytogenetic studies showed that the tumor was of nonhost origin suggesting it was derived from the transplanted neural stem cells. Microsatellite and HLA analysis demonstrated that the tumor is derived from at least two donors. CONCLUSIONS: This is the first report of a human brain tumor complicating neural stem cell therapy. The findings here suggest that neuronal stem/progenitor cells may be involved in gliomagenesis and provide the first example of a donor-derived brain tumor. Further work is urgently needed to assess the safety of these therapies.

  12. Identification of 4 ataxia telangiectasia cell lines hypersensitive to. gamma. -irradiation but not to hydrogen peroxide

    Energy Technology Data Exchange (ETDEWEB)

    Cantoni, O.; Sestili, P.; Santoro, M.P.; Tannoia, M.C.; Cattabeni, F. (Universita degli Studi di Urbino (Italy). Istituto di Farmacologia e Farmacognosia and Centro di Farmacologia Oncologia Sperimentale); Novelli, G.; Dallapiccola, B. (Universit degli Studi di Urbino (Italy). Cattedra di Genetica); Fiorilli, M. (Universita di Roma ' La Sapienze' (Italy). Cattedra di Allergologia e Immunologia Clinica)

    1989-09-01

    The effct of hydrogen peroxide on the rate of semi-conservative DNA synthesis in ataxia telangiectasia (AT) and normal human lymphoblastoid cells was investigated. The rate of DNA synthesis in AT cells was not depressed to a lesser extent than in normal cells, as might have been expected since H{sub 2O2} is a radiomimetic agent. On the contrary, 4 AT cell lines displayed a higher sensitivity to the inhibitory effect of H{sub 2O2} on DNA synthesis than 2 normal cell lines. Comparable levels of cytotoxicity were detected in cell vaibility studies. Furthermore, neither the level of DNA breakage produced by H{sub 2O2}, nor the rate of repair of these lesions was signigicantly different in normal and AT cells. Together, these results indicate that the AT cell lines utilized in this study are not hypersensitive to the oxidant. It is suggested that H-2-O-2 may not induce lethality via the direct ation of the hydroxyl radical (OH). (Author). 20 refs.; 3 figs.; 1 tab.

  13. Ataxia-telangiectasia mutated (ATM) deficiency decreases reprogramming efficiency and leads to genomic instability in iPS cells

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    Kinoshita, Taisuke [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Nagamatsu, Go, E-mail: gonag@sc.itc.keio.ac.jp [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012 (Japan); Kosaka, Takeo [Department of Urology, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Takubo, Keiyo [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan); Hotta, Akitsu [Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012 (Japan); Department of Reprogramming Science, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto (Japan); Ellis, James [Ontario Human iPS Cell Facility, Molecular Genetics, University of Toronto, Developmental and Stem Cell Biology, SickKids, Toronto, Canada MG1L7 (Canada); Suda, Toshio, E-mail: sudato@sc.itc.keio.ac.jp [Department of Cell Differentiation, The Sakaguchi Laboratory, School of Medicine, Keio University, Tokyo 160-8582 (Japan)

    2011-04-08

    Highlights: {yields} iPS cells were induced with a fluorescence monitoring system. {yields} ATM-deficient tail-tip fibroblasts exhibited quite a low reprogramming efficiency. {yields} iPS cells obtained from ATM-deficient cells had pluripotent cell characteristics. {yields} ATM-deficient iPS cells had abnormal chromosomes, which were accumulated in culture. -- Abstract: During cell division, one of the major features of somatic cell reprogramming by defined factors, cells are potentially exposed to DNA damage. Inactivation of the tumor suppressor gene p53 raised reprogramming efficiency but resulted in an increased number of abnormal chromosomes in established iPS cells. Ataxia-telangiectasia mutated (ATM), which is critical in the cellular response to DNA double-strand breaks, may also play an important role during reprogramming. To clarify the function of ATM in somatic cell reprogramming, we investigated reprogramming in ATM-deficient (ATM-KO) tail-tip fibroblasts (TTFs). Although reprogramming efficiency was greatly reduced in ATM-KO TTFs, ATM-KO iPS cells were successfully generated and showed the same proliferation activity as WT iPS cells. ATM-KO iPS cells had a gene expression profile similar to ES cells and WT iPS cells, and had the capacity to differentiate into all three germ layers. On the other hand, ATM-KO iPS cells accumulated abnormal genome structures upon continuous passages. Even with the abnormal karyotype, ATM-KO iPS cells retained pluripotent cell characteristics for at least 20 passages. These data indicate that ATM does participate in the reprogramming process, although its role is not essential.

  14. Radiation Dose-effects on Cell Cycle, Apoptosis, and Marker Expression of Ataxia Telangiectasia-Heterozygous Human Breast Epithelial Cells

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    Cruz, A.; Bors, K.; Jansen, H.; Richmond, R.

    2003-01-01

    Ataxia-telangiectasia (A-T) is a radiation-sensitive genetic condition. AT-heterozygous human mammary epithelial cells (HMEC) were irradiated using a Cs137 source in order to compare cell cycle, apoptosis, and marker expression responses across 3 radiation doses. No differences in cell cycle and apoptosis were found with any of the radiation doses used (30, 60, and 90 rads) compared with the unirradiated control (0 rad). At the same doses, however, differences were found in marker expression, such as keratin 18 (kl8), keratin 14 (k14), insulin-like growth factor I receptor (IGF-IR), and connexin 43 (cx43). This may indicate that radiation sensitivity in the heterozygous state may be initiated through signal transduction responses.

  15. Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis

    Directory of Open Access Journals (Sweden)

    Zhe-Wei Zhang

    2015-01-01

    Full Text Available Background: Caffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR. Methods: Immunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine. A mouse xenograft model was used to study the effects of caffeine on the DNA damage responses. Western blotting was used to investigate the effects of caffeine pretreatment on the ATM-Chk2-p53-Puma axis, while real-time polymerase chain reaction (RT-PCR assessed changes in messenger RNA levels of p53 and downstream targets responding to IR. Finally, terminal deoxynucleotidyl transferase-dUTP nick end labeling assay. Western blotting and colony formation assay were used to measure the effects of caffeine on radiation-related apoptosis. All of the data were analyzed with a two-tailed Student′s t-test. Results: Immunofluorescent staining showed that caffeine pretreatment profoundly suppressed the formation of γH2AXand p53-binding protein 1 foci in RT4 cells in response to irradiation. Cellular and animal experiments suggested that this suppression was mediated by suppression of the ATM-Chk2-p53-Puma DNA damage-signaling axis. RT-PCR indicated caffeine also attenuated transactivation of p53 and p53-inducible genes. The colony formation assay revealed that caffeine displayed radioprotective effects on RT4 cells in response to low-dose radiation compared to the radiosensitization effects on T24 cells. Conclusion: Caffeine may inhibit IR-related apoptosis of bladder cancer RT4 cells by suppressing activation of the ATM-Chk2-p53-Puma axis.

  16. Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis.

    Science.gov (United States)

    Zhang, Zhe-Wei; Xiao, Jing; Luo, Wei; Wang, Bo-Han; Chen, Ji-Min

    2015-11-05

    Caffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM) activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR). Immunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine. A mouse xenograft model was used to study the effects of caffeine on the DNA damage responses. Western blotting was used to investigate the effects of caffeine pretreatment on the ATM-Chk2-p53-Puma axis, while real-time polymerase chain reaction (RT-PCR) assessed changes in messenger RNA levels of p53 and downstream targets responding to IR. Finally, terminal deoxynucleotidyl transferase-dUTP nick end labeling assay. Western blotting and colony formation assay were used to measure the effects of caffeine on radiation-related apoptosis. All of the data were analyzed with a two-tailed Student's t-test. Immunofluorescent staining showed that caffeine pretreatment profoundly suppressed the formation of γH2AXand p53-binding protein 1 foci in RT4 cells in response to irradiation. Cellular and animal experiments suggested that this suppression was mediated by suppression of the ATM-Chk2-p53-Puma DNA damage-signaling axis. RT-PCR indicated caffeine also attenuated transactivation of p53 and p53-inducible genes. The colony formation assay revealed that caffeine displayed radioprotective effects on RT4 cells in response to low-dose radiation compared to the radiosensitization effects on T24 cells. Caffeine may inhibit IR-related apoptosis of bladder cancer RT4 cells by suppressing activation of the ATM-Chk2-p53-Puma axis.

  17. Establishment of immortal normal and ataxia telangiectasia fibroblast cell lines by introduction of the hTERT gene

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    Nakamura, Hideaki; Fukami, Hiroko; Hayashi, Yuko; Kiyono, Tohru; Ishizaki, Kanji [Aichi Cancer Center, Nagoya (Japan). Research Inst.; Nakatsugawa, Shigekazu; Hamaguchi, Michinari [Nagoya Univ. (Japan). School of Medicine

    2002-06-01

    To establish immortal human cells, we introduced the human catalytic subunit of telomerase (hTERT) gene into skin fibroblast cells obtained from normal and ataxia telangiectasia (AT) individuals of Japanese origin. After hTERT introduction, these cells continue to grow beyond a population doubling number of 200 while maintaining their original radiosensitivity. Inductions of p53, phosphorylation of Serl5 in p53, and induction of p21 by X-ray irradiation in immortal cells derived from normal individual were not affected by the hTERT introduction. Both normal and AT immortal cells exhibited an apparent inhibition of growth as original primary cells when they reached confluence. Karyotype analysis has revealed that they are in a diploid range. These results suggest that cells immortalized by hTERT introduction retain their original characteristics except for immortalization, and that they may be useful for analyzing various effects of radiation on human cells. (author)

  18. Ataxia-Telangiectasia

    OpenAIRE

    J Gordon Millichap

    1990-01-01

    São apresentados os casos de dois irmãos com ataxia-telangiectasia, estudados sob os pontos de vista clínico, eletrencefalográfico, liquórico e encefalográfico. O autor resume os achados de diversos autores e chama a atenção para a regressão parcial da síndrome cerebelar em ambos os pacientes, fato ainda não referido na literatura.

  19. Genetics Home Reference: ataxia-telangiectasia

    Science.gov (United States)

    ... Email Facebook Twitter Home Health Conditions Ataxia-telangiectasia Ataxia-telangiectasia Printable PDF Open All Close All Enable Javascript to view the expand/collapse boxes. Description Ataxia-telangiectasia is a rare inherited disorder that affects ...

  20. Biological and molecular mechanisms of sulfur mustard analogue-induced toxicity in JB6 and HaCaT cells: possible role of ataxia telangiectasia-mutated/ataxia telangiectasia-Rad3-related cell cycle checkpoint pathway.

    Science.gov (United States)

    Tewari-Singh, Neera; Gu, Mallikarjuna; Agarwal, Chapla; White, Carl W; Agarwal, Rajesh

    2010-06-21

    Effective medical treatment and preventive measures for chemical warfare agent sulfur mustard (HD)-caused incapacitating skin toxicity are lacking, because of limited knowledge of its mechanism of action. The proliferating basal epidermal cells are primary major sites of attack during HD-caused skin injury. Therefore, employing mouse JB6 and human HaCaT epidermal cells, here, we investigated the molecular mechanism of HD analogue 2-chloroethyl ethyl sulfide (CEES)-induced skin cytotoxicity. As compared to the control, up to 1 mM CEES treatment of these cells for 2, 4, and 24 h caused dose-dependent decreases in cell viability and proliferation as measured by DNA synthesis, together with S and G2-M phase arrest in cell cycle progression. Mechanistic studies showed phosphorylation of DNA damage sensors and checkpoint kinases, ataxia telangiectasia-mutated (ATM) at ser1981 and ataxia telangiectasia-Rad3-related (ATR) at ser428 within 30 min of CEES exposure, and modulation of S and G2-M phase-associated cell cycle regulatory proteins, which are downstream targets of ATM and ATR kinases. Hoechst-propidium iodide staining demonstrated that CEES-induced cell death was both necrotic and apoptotic in nature, and the latter was induced at 4 and 24 h of CEES treatment in HaCaT and JB6 cells, respectively. An increase in caspase-3 activity and both caspase-3 and poly(ADP-ribose)polymerase (PARP) cleavage coinciding with CEES-caused apoptosis in both cell lines suggested the involvement of the caspase pathway. Together, our findings suggest a DNA-damaging effect of CEES that activates ATM/ATR cell cycle checkpoint signaling as well as caspase-PARP pathways, leading to cell cycle arrest and apoptosis/necrosis in both JB6 and HaCaT cells. The identified molecular targets, quantitative biomarkers, and epidermal cell models in this study have the potential and usefulness in rapid development of effective prophylactic and therapeutic interventions against HD-induced skin toxicity.

  1. What Is Ataxia-Telangiectasia?

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    ... About A-T Research Fundraising About Us About Ataxia-telangiectasia About A-T » WHAT IS A- ... develop slurred or distorted speech, and swallowing problems. Ataxia... The onset of this ataxia marks the beginning ...

  2. Ataxia-telangiectasia: future prospects

    Directory of Open Access Journals (Sweden)

    Chaudhary MW

    2014-09-01

    Full Text Available Mohammed Wajid Chaudhary, Raidah Saleem Al-Baradie Pediatric Neurology, Neurosciences Centre, King Fahad Specialist Hospital, Dammam, Kingdom of Saudi Arabia Abstract: Ataxia-telangiectasia (A-T is an autosomal recessive multi-system disorder caused by mutation in the ataxia-telangiectasia mutated gene (ATM. ATM is a large serine/threonine protein kinase, a member of the phosphoinositide 3-kinase-related protein kinase (PIKK family whose best-studied function is as master controller of signal transduction for the DNA damage response (DDR in the event of double strand breaks (DSBs. The DDR rapidly recognizes DNA lesions and initiates the appropriate cellular programs to maintain genome integrity. This includes the coordination of cell-cycle checkpoints, transcription, translation, DNA repair, metabolism, and cell fate decisions, such as apoptosis or senescence. DSBs can be generated by exposure to ionizing radiation (IR or various chemical compounds, such as topoisomerase inhibitors, or can be part of programmed generation and repair of DSBs via cellular enzymes needed for the generation of the antibody repertoire as well as the maturation of germ cells. AT patients have immunodeficiency, and are sterile with gonadal dysgenesis as a result of defect in meiotic recombination. In the cells of nervous system ATM has additional role in vesicle dynamics as well as in the maintenance of the epigenetic code of histone modifications. Moderate levels of ATM are associated with prolonged lifespan through resistance to oxidative stress. ATM inhibitors are being viewed as potential radiosensitizers as part of cancer radiotherapy. Though there is no cure for the disease at present, glucocorticoids have been shown to induce alternate splicing site in the gene for ATM partly restoring its activity, but their most effective timing in the disease natural history is not yet known. Gene therapy is promising but large size of the gene makes it technically difficult

  3. Targeting the Ataxia Telangiectasia Mutated-null phenotype in chronic lymphocytic leukemia with pro-oxidants

    Science.gov (United States)

    Agathanggelou, Angelo; Weston, Victoria J.; Perry, Tracey; Davies, Nicholas J.; Skowronska, Anna; Payne, Daniel T.; Fossey, John S.; Oldreive, Ceri E.; Wei, Wenbin; Pratt, Guy; Parry, Helen; Oscier, David; Coles, Steve J.; Hole, Paul S.; Darley, Richard L.; McMahon, Michael; Hayes, John D.; Moss, Paul; Stewart, Grant S.; Taylor, A. Malcolm R.; Stankovic, Tatjana

    2015-01-01

    Inactivation of the Ataxia Telangiectasia Mutated gene in chronic lymphocytic leukemia results in resistance to p53-dependent apoptosis and inferior responses to treatment with DNA damaging agents. Hence, p53-independent strategies are required to target Ataxia Telangiectasia Mutated-deficient chronic lymphocytic leukemia. As Ataxia Telangiectasia Mutated has been implicated in redox homeostasis, we investigated the effect of the Ataxia Telangiectasia Mutated-null chronic lymphocytic leukemia genotype on cellular responses to oxidative stress with a view to therapeutic targeting. We found that in comparison to Ataxia Telangiectasia Mutated-wild type chronic lymphocytic leukemia, pro-oxidant treatment of Ataxia Telangiectasia Mutated-null cells led to reduced binding of NF-E2 p45-related factor-2 to antioxidant response elements and thus decreased expression of target genes. Furthermore, Ataxia Telangiectasia Mutated-null chronic lymphocytic leukemia cells contained lower levels of antioxidants and elevated mitochondrial reactive oxygen species. Consequently, Ataxia Telangiectasia Mutated-null chronic lymphocytic leukemia, but not tumors with 11q deletion or TP53 mutations, exhibited differentially increased sensitivity to pro-oxidants both in vitro and in vivo. We found that cell death was mediated by a p53- and caspase-independent mechanism associated with apoptosis inducing factor activity. Together, these data suggest that defective redox-homeostasis represents an attractive therapeutic target for Ataxia Telangiectasia Mutated-null chronic lymphocytic leukemia. PMID:25840602

  4. Nitric Oxide Induces Ataxia Telangiectasia Mutated (ATM) Protein-dependent γH2AX Protein Formation in Pancreatic β Cells*

    Science.gov (United States)

    Oleson, Bryndon J.; Broniowska, Katarzyna A.; Schreiber, Katherine H.; Tarakanova, Vera L.; Corbett, John A.

    2014-01-01

    In this study, the effects of cytokines on the activation of the DNA double strand break repair factors histone H2AX (H2AX) and ataxia telangiectasia mutated (ATM) were examined in pancreatic β cells. We show that cytokines stimulate H2AX phosphorylation (γH2AX formation) in rat islets and insulinoma cells in a nitric oxide- and ATM-dependent manner. In contrast to the well documented role of ATM in DNA repair, ATM does not appear to participate in the repair of nitric oxide-induced DNA damage. Instead, nitric oxide-induced γH2AX formation correlates temporally with the onset of irreversible DNA damage and the induction of apoptosis. Furthermore, inhibition of ATM attenuates cytokine-induced caspase activation. These findings show that the formation of DNA double strand breaks correlates with ATM activation, irreversible DNA damage, and ATM-dependent induction of apoptosis in cytokine-treated β cells. PMID:24610783

  5. [Jaridonin, a new diterpenoid from Isodon rubescens, induces cell cycle arrest in gastric cancer cells through activating ataxia telangiectasia mutated kinase].

    Science.gov (United States)

    Ma, Y C; Su, N; Zhao, N M; Li, Q Y; Zhang, M; Zhao, H W; Liu, H M; Qin, Y H

    2016-04-01

    To study the effects of Jaridonin, a novel diterpenoid from isodon rubescens, on the cell cycle of human gastric cancer cells and its molecular mechanism of action. Flow cytometry was used to analyze the cell cycle distribution and expression of ataxia telangiectasia mutated kinase (ATM) after Jaridonin treatment. Western blot was performed to detect the expression of cell cycle-related proteins. The results of flow cytometry showed that the percentages of MGC-803 cells in G(2)/M phase at 6 hours after 0, 10, 20 μmol/L Jaridonin-treatment were (10.8±2.2)%, (18.2±2.5)%, (27.3±3.2)%, respectively; those at 12 hours after Jaridonin-treatment were (12.0±1.5)%, (24.1±2.0)% and (39.7±5.2)%, respectively, indicating a G2/M phase arrest of MGC-803 cells was resulted in a time- and dose-dependent manner. The expressions of ATM, Chk1, Chk2, phosphorylated Cdc2 and CDK2 were up-regulated in the MGC-803 cells after Jaridonin treatment, while the levels of Cdc2 and CDK2 were decreased. KU-55933, an inhibitor of ATM, reversed the expression of relevant proteins and G(2)/M phase arrest induced by Jaridonin. Jaridonin can significantly induce G(2)/M arrest in gastric cancer MGC-803 cells. Its mechanism may be related to the activation of ATM and Chk1/2, and inactivation of Cdc2 and CDK2 phosphorylation.

  6. Spontaneous chromosomal aberrations in Fanconi anaemia, ataxia telangiectasia fibroblast and Bloom's syndrome lymphoblastoid cell lines as detected by conventional cytogenetic analysis and fluorescence in situ hybridisation (FISH) technique.

    Science.gov (United States)

    Sakamoto Hojo, E T; van Diemen, P C; Darroudi, F; Natarajan, A T

    1995-02-01

    Several primary and transformed human cell lines derived from cancer prone patients are employed routinely for biochemical and DNA repair studies. Since transformation leads to some chromosomal instability a cytogenetic analysis of spontaneous chromosome aberrations in fibroblast cell lines derived from patients with Fanconi anaemia (FA), ataxia telangiectasia (AT), and in lymphoblastoid cell lines derived from patients with Bloom's syndrome (BS), was undertaken. Unstable aberrations were analysed in Giemsa stained preparations and the chromosome painting technique was used for evaluating the frequencies of stable aberrations (translocations). In addition, the frequency of sister-chromatid exchanges (SCEs) was determined in differentially stained metaphases. The SV40-transformed fibroblasts from these cell lines have higher frequencies of unstable aberrations than the primary fibroblasts. In the four lymphoblastoid cell lines derived from BS patients higher frequencies of spontaneously occurring chromosomal aberrations in comparison to normal TK6wt cells were also evident. The frequency of spontaneously occurring chromosome translocations was determined with fluorescence in situ hybridisation (FISH) and using DNA libraries specific for chromosomes 1, 2, 3, 4, 7, 8, 11, 14, 19, 20 and X. The translocation levels were found to be elevated for primary FA fibroblasts and lymphoblastoid cells derived from BS patients in comparison with control cell lines, hetero- and homozygote BS cell lines not differing in this respect. The SV40-transformed cell lines showed very high frequencies of translocations independent of their origin and almost every cell contained at least one translocation. In addition, clonal translocations were found in transformed control TK6wt and AT cell lines for chromosomes 20 and 14, respectively. The spontaneous frequencies of SCEs were similar in transformed fibroblasts derived from normal individuals and AT patients, whereas in SV40-transformed FA

  7. MicroRNA-223 Enhances Radiation Sensitivity of U87MG Cells In Vitro and In Vivo by Targeting Ataxia Telangiectasia Mutated

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Liping; Zhu, Ji [Departments of Radiation Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Zaorsky, Nicholas G. [Department of Radiation Oncology, Fox Chase Cancer Center, Philadelphia, Pennsylvania (United States); Deng, Yun [Departments of Radiation Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Wu, Xingzhong [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China); Liu, Yong [Departments of Radiation Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Liu, Fangqi; Cai, Guoxiang; Gu, Weilie [Department of Colorectal Cancer, Fudan University, Shanghai Cancer Center, Shanghai (China); Shen, Lijun [Departments of Radiation Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Zhang, Zhen, E-mail: zhenzhang6@hotmail.com [Departments of Radiation Oncology, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China)

    2014-03-15

    Purpose: Ataxia telangiectasia mutated (ATM) protein is important in the DNA damage response because it repairs radiation-induced damage in cancers. We examined the effect of microRNA-223 (miR-223), a regulator of ATM expression, on radiation sensitivity of cancer cells. Methods and Materials: Human embryonic kidney 293 T (293T) cells were infected with pLL3.7-miR-223 plasmid to generate the pLL3.7-miR-223 and -empty virus (EV) lentivirus (miR-223 and EV). A dual luciferase assay in which the reporter contained wild-type 3′ untranslated region (UTR) of ATM was performed. U87MG cells were infected with miR-223 or EV to establish the overexpressed stable cell lines (U87-223 or U87-EV, respectively). Cells were irradiated in vitro, and dose enhancement ratios at 2 Gy (DER{sub 2}) were calculated. Hind legs of BALB/c athymic mice were injected with U87-223 or U87-EV cells; after 2 weeks, half of the tumors were irradiated. Tumor volumes were tracked for a total of 5 weeks. Results: The dual luciferase reporter assay showed a significant reduction in luciferase activity of 293T cells cotransfected with miR-223 and the ATM 3′UTR compared to that in EV control. Overexpression of miR-223 in U87MG cells showed that ATM expression was significantly downregulated in the U87-223 cells compared to that in U87-EV (ATM/β-actin mRNA 1.0 vs 1.5, P<.05). U87-223 cells were hypersensitive to radiation compared to U87-EV cells in vitro (DER{sub 2} = 1.32, P<.01). Mice injected with miR-223-expressing tumors had almost the same tumors after 3 weeks (1.5 cm{sup 3} vs 1.7 cm{sup 3}). However, irradiation significantly decreased tumor size in miR-223-expressing tumors compared to those in controls (0.033 cm{sup 3} vs 0.829 cm{sup 3}). Conclusions: miR-223 overexpression downregulates ATM expression and sensitizes U87 cells to radiation in vitro and in vivo. MicroRNA-223 may be a novel cancer-targeting therapy, although its cancer- and patient-specific roles are

  8. T-cell ALL in ataxia telangiectasia cured with only 7 weeks of anti-leukemic therapy

    DEFF Research Database (Denmark)

    Hersby, Ditte S; Sehested, Astrid; Kristensen, Kim

    2015-01-01

    A 20-month-old girl diagnosed with T-cell acute lymphoblastic leukemia was treated according to the Nordic NOPHO ALL2000 protocol. The patient developed severe immunosuppression and experienced life-threatening adenovirus infection, which was treated with ribavirin and cidofovir. α-fetoprotein wa......A 20-month-old girl diagnosed with T-cell acute lymphoblastic leukemia was treated according to the Nordic NOPHO ALL2000 protocol. The patient developed severe immunosuppression and experienced life-threatening adenovirus infection, which was treated with ribavirin and cidofovir. α...

  9. [Ataxia telangiectasia: review of 13 new cases].

    Science.gov (United States)

    Valbuena, O; Póo, P; Campistol, J; Vernet, A; Fernández-Alvarez, E; Sierra, I; Gean, E

    1996-01-01

    We report the review of 13 patients who were diagnosed of ataxia telangiectasia before 6 years of age. All of them manifested cerebelous ataxia, oculocutaneus telangiectasias (11), sinopulmonary infections (9), dystonia (9), oculomotor apraxia (9) and Burkitt linfoma (1). We analyse the most common presentation of the disease in early stages and the complementary studies performed. The prompt diagnosis allow us a better control of infections, malignant process and finally the possibility of genetic counseling.

  10. The Functional Role of the Ataxia Telangiectasia Gene

    National Research Council Canada - National Science Library

    Gautier, Jean

    2000-01-01

    Ataxia Telangiectasia (A-T) is an autosomal recessive disease characterized by a progressive cerebellar ataxia, severe immune deficiencies, gonadal atrophy, telangiectases, increased risk for cancer, particularly lymphomas...

  11. The Functional Role of the Ataxia Telangiectasia Gene

    National Research Council Canada - National Science Library

    Gautier, Jean

    1999-01-01

    Ataxia Telangiectasia (A-T) is an autosomal recessive disease characterized by a progressive cerebellar ataxia, severe immune deficiencies, gonadal atrophy, telangiectases, increased risk for cancer, particularly lymphomas...

  12. Anti-oxidative capacity in patients with ataxia telangiectasia

    Science.gov (United States)

    Reichenbach, J; Schubert, R; Schwan, C; Müller, K; Böhles, H J; Zielen, S

    1999-01-01

    Highly reactive oxygen species (ROS) are involved in T-cell activation and in the defense against environmental pathogens. An imbalance of ROS generation and detoxifying scavenger enzymes could contribute to the increased susceptibility to cancer and infections in ataxia telangiectasia. We studied oxidative status, i.e. plasma total antioxidant capacity (TEAC), retinol, α-tocopherol, ubiquinol, and the number of activated T cells in 10 patients with ataxia telangiectasia (AT) compared to age-matched healthy controls. As expected, patients showed significantly increased levels of activated human leukocyte antigen-DR and CD45RO expressing T cells. TEAC levels as well as the exogenous antioxidants retinol and α-tocopherol were significantly reduced in patients. In addition, patients showed slightly reduced plasma levels of the endogenous ROS scavenger enzyme ubiquinol (Q10). Although no correlation between number of activated T-cells and antioxidant capacity could be demonstrated, an increase in ROS and a diminished reactive oxygen scavenger capacity may be involved in the disease process of patients with AT. PMID:10469059

  13. Ataxia Telangiectasia - A Report of a case in Port Harcourt

    African Journals Online (AJOL)

    TNHJOURNALPH

    Ataxia Telangiectasia - A Report of a case in Port Harcourt. Lucy Yaguo-Ide, Tochi Uchenwa, BalafamaAlex-Hart, Alice Nte, Chidi Ezeani. Department ofPeadiatrics and Child Health, University of Port Harcourt Teaching Hospital, Port. Harcourt, Nigeria. ABSTRACT. BACKGROUND. Ataxia telangiectasia is acomplex multi-.

  14. DNA synthesis in ataxia telangiectasia

    OpenAIRE

    Jaspers, Nicolaas

    1985-01-01

    textabstractAfter the discovery that cultured cells from AT patients are hypersensitive to ionizing radiation the suggestion was made that AT-could be the 1 X-ray-analogue 1 of xeroderma pigmentosum. The latter syndrome (XP) is characterized by hypersensitivity to short-wave UV-radiation, caused by a reduced ability to properly remove UV-induced DNA damage. The evidence for a DNA repair defect in AT cells is not as strong as in the case of XP (see section 2.2.5 of this thesis). Different XP p...

  15. DNA synthesis in ataxia telangiectasia

    NARCIS (Netherlands)

    N.G.J. Jaspers (Nicolaas)

    1985-01-01

    textabstractAfter the discovery that cultured cells from AT patients are hypersensitive to ionizing radiation the suggestion was made that AT-could be the 1 X-ray-analogue 1 of xeroderma pigmentosum. The latter syndrome (XP) is characterized by hypersensitivity to short-wave UV-radiation, caused by

  16. Accurate diagnostics of ataxia-telangiectasia cellular phenotype by employing in vitro lymphocyte radiosensitivity testing

    Directory of Open Access Journals (Sweden)

    Vujić Dragana S.

    2013-01-01

    Full Text Available In this paper we present the data of lymphocyte radiosensitivity testing used for characterization of radiosensitive cellular phenotype and diagnostics of ataxia-telangiectasia disease. We point out the advantage of lymphocyte micronucleus test (CBMN over other cellular tests for assessment of radiosensitivity: the first advantage of CBMN is that primary patient cells are used (less than 1 ml, the second one is that the results of testing are obtained within 3 days and there is no need for establishing a patient-derived cell line, which requires additional time and application of more expensive methods. The third advantage of CBMN method is that it gives information about proliferative ability of cells, which can recognize dysfunctional ataxia-telangiectasia mutated protein. The results are fast and accurate in diagnostics of ataxia-telagiectasia diseases.

  17. Ataxia telangiectasia mutated inhibits oxidative stress-induced apoptosis by regulating heme oxygenase-1 expression.

    Science.gov (United States)

    Yu, Ji Hoon; Cho, Soon Ok; Lim, Joo Weon; Kim, Nanhee; Kim, Hyeyoung

    2015-03-01

    Ataxia telangiectasia (AT) is caused by mutational inactivation of the ataxia telangiectasia mutated (Atm) gene, which is involved in DNA repair. Increased oxidative stress has been shown in human AT cells and neuronal tissues of Atm-deficient mice. Heme oxygenase-1 (HO-1) is an inducible antioxidant enzyme and protects cells against oxidative stress. The purpose of this study is to determine whether ATM induces antioxidant enzyme HO-1 and protects cells from oxidative stress-mediated apoptosis by driving the activation of PKC-δ and NF-κB, by increasing cell viability, and by downregulating DNA fragmentation and apoptotic indicators (apoptosis-inducing factor and cleaved caspase-3). AT fibroblasts stably transfected with human full-length ATM cDNA (YZ5 cells) or the empty vector (MOCK cells) were treated with H2O2 as a source of reactive oxygen species (ROS). As a result, transfection with ATM inhibited ROS-induced cell death and DNA fragmentation in MOCK cells. Transfection with ATM induced expression of HO-1 which was mediated by PKC-δ and NF-κB in H2O2-treated MOCK cells. ZnPP, an HO-1 inhibitor, and transfection with HO-1 siRNA increased ROS levels and apoptosis, whereas hemin, an HO-1 activator, reduced ROS levels and apoptosis in H2O2-treated YZ5 cells. Rottlerin, a PKC-δ inhibitor, inhibited NF-κB activation and HO-1 expression in H2O2-treated YZ5 cells. MOCK cells showed increased cell death, DNA fragmentation, and apoptotic indicators compared to YZ5 cells exposed to H2O2. In addition, transfection with p65 siRNA increased ROS levels and DNA fragmentation, but decreased HO-1 protein levels in H2O2-treated YZ5 cells. In conclusion, ATM induces HO-1 expression via activation of PKC-δ and NF-κB and inhibits oxidative stress-induced apoptosis. A loss of HO-1 induction may explain why AT patients are vulnerable to oxidative stress. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Síndrome de Ataxia-Telangiectasia

    Directory of Open Access Journals (Sweden)

    Amauri Batista da Silva

    1971-06-01

    Full Text Available A ataxia-telangiectasia, doença de Mme. Louis-Bar, é caracterizada pela associação de ataxia cerebelar progressiva, em geral com início na primeira infância, telangiectasas óculo-cutâneas, movimentos coreoatetósicos, tendência a infecções repetidas do sistema respiratório, retardo estaturo-ponderal, demenciação. São mais ou menos freqüentes os tumores do sistema reticuloendotelial. A doença é geralmente familiar, transmitida por genes recessivos, autossômicos, não ligados ao sexo. A alteração bioquímica mais encontrada consiste na diminuição ou ausência completa da fração A das gamaglobulinas, bem como na perturbação das reações de hipersensibilidade retardada. Os AA. relatam o estudo clínico, biológico e pneumencefalográfico de uma criança de 3 anos de idade, apresentando essa enfermidade desde os 18 meses de vida, sem antecedentes familiares.

  19. Cranial MRI in ataxia-telangiectasia

    Energy Technology Data Exchange (ETDEWEB)

    Sardanelli, F. [Dept. of Radiology, Univ. of Genoa (Italy); Parodi, R.C. [Dept. of Radiology, Univ. of Genoa (Italy); Ottonello, C. [Dept. of Radiology, Univ. of Genoa (Italy); Renzetti, P. [Dept. of Radiology, Univ. of Genoa (Italy); Saitta, S. [Dept. of Radiology, Univ. of Genoa (Italy); Lignana, E. [G. Gaslini Inst., Genoa (Italy); Mancardi, G.L. [Dept. of Neurology, Univ. of Genoa (Italy)

    1995-01-01

    We examined five males with laboratory-confirmed ataxia-telangiectasia (AT), aged 9-28 years, several times by MRI (9 examinations: 5 at 0.15 T, 3 at 0.5 T, 1 at 1.5 T). Intermediate, T1-, T2- and T2{sup *}-weighted spin-echo and gradient-echo sequences were performed. All patients showed vermian atrophy, enlarged fourth ventricle and cisterna magna; four showed cerebellar hemisphere atrophy; two enlarged infracerebellar subarachnoid spaces and four patients had sinusitis. No focal areas of abnormal signal were seen in the brain, diffuse high signal was found in the central cerebral white matter of the oldest patient. AT is an important human model of inherited cancer susceptibility and multisystem ageing; as in xeroderma pigmentosum and other ``breakage syndromes``, ionising radiation should be avoided. When imaging is necessary, MRI should be preferred to CT in patients known or suspected to have AT and those with undefined paediatric ataxias of nontraumatic origin. If atrophy of only the cerebellum, especially the vermis, is noted, laboratory research should be performed to confirm the diagnosis of AT. (orig.)

  20. [Ataxia telangiectasia: what impact in clinical oncology?].

    Science.gov (United States)

    Stoppa-Lyonnet, D; Aurias, A

    1992-01-01

    Ataxia telangiectasia (AT) is a hereditary disease transmitted in a recessive mode and characterized by chromosomal instability and radiosensitivity. AT patients have a 100-fold higher risk of cancer than the general population. Although AT is a rare disease of which the frequency has been estimated to be 1/40,000, the frequency of the heterozygosity status, when assessed with the Hardy-Weinberg equation is high (about 1.4%). Parents of AT children, thus obligate AT carriers, show chromosomal instability and radiosensitivity, but at a lower level than AT patients. Assuming that these AT characteristics deal with the cancer predisposition, it can be hypothesized that AT heterozygote individuals have a higher cancer susceptibility than the general population. To test this hypothesis, M Swift's group compared cancer incidence rates from adult blood relatives of AT patients with controls. The risk of cancer in AT heterozygotes could be increased by 3.5 and, for carrier women, the breast cancer risk could be increased by 5.1. Actually, the diagnosis of the AT heterozygote status is not possible. However, the near cloning of the gene (or genes) for the disease will permit to identify the AT carriers in a population of patients suffering from cancer and to assess precisely the impact of AT heterozygosity in the genetic predisposition to cancer.

  1. Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis

    National Research Council Canada - National Science Library

    Zhe-Wei Zhang Jing Xiao Wei Luo Bo-Han Wang Ji-Min Chen

    2015-01-01

    ...; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR). Methods...

  2. Neurodegeneration in ataxia-telangiectasia is caused by horror autotoxicus.

    Science.gov (United States)

    Kuljis, R O; Aguila, M C

    1999-05-01

    Ataxia-telangiectasia (A-T) is a pleiotropic, multi-system disorder with manifestations that include immune deficiency, sensitivity to ionizing radiation and neoplasms. Many of these manifestations are understood in principle since the identification in A-T patients of mutations in a gene encoding a protein kinase that plays a key role in signaling and repair of DNA damage. However, the cause of the neurodegeneration that afflicts patients with A-T for at least a decade before they succumb to overwhelming infections or malignancy remains mysterious. Based on our work in a mouse model of A-T and previous evidence of extra-neural autoimmune disorders in A-T, we postulate that the neurodegenerative process in A-T is not due to a function for A-T mutated (ATM) essential for the postnatal brain, but to an autoimmune process (hence 'horror autotoxicus', Paul Ehrlich's term for autoimmune disorder). This hypothetical mechanism may be analogous to that in the so-called 'paraneoplastic' neurodegenerative syndromes in patients with various malignancies. Thus, alterations in the balance between cellular and humoral immunity in A-T probably result in autoantibodies to cerebral epitopes shared with cells of the immune system. This hypothesis has important implications for the understanding and development of effective palliative and even preventative strategies for A-T, and probably for other so far relentlessly progressive neurodegenerative disorders.

  3. Altered corticomotor-cerebellar integrity in young ataxia telangiectasia patients.

    Science.gov (United States)

    Sahama, Ishani; Sinclair, Kate; Fiori, Simona; Pannek, Kerstin; Lavin, Martin; Rose, Stephen

    2014-09-01

    Magnetic resonance imaging (MRI) research in identifying altered brain structure and function in ataxia-telangiectasia, an autosomal recessive neurodegenerative disorder, is limited. Diffusion-weighted MRI were obtained from 11 ataxia telangiectasia patients (age range, 7-22 years; mean, 12 years) and 11 typically developing age-matched participants (age range, 8-23 years; mean, 13 years). Gray matter volume alterations in patients were compared with those of healthy controls using voxel-based morphometry, whereas tract-based spatial statistics was employed to elucidate white matter microstructure differences between groups. White matter microstructure was probed using quantitative fractional anisotropy and mean diffusivity measures. Reduced gray matter volume in both cerebellar hemispheres and in the precentral-postcentral gyrus in the left cerebral hemisphere was observed in ataxia telangiectasia patients compared with controls (P cerebellar hemispheres, anterior/posterior horns of the medulla, cerebral peduncles, and internal capsule white matter, particularly in the left posterior limb of the internal capsule and corona radiata in the left cerebral hemisphere, was observed in patients compared with controls (P cerebellar hemisphere and the white matter of the superior lobule of the right cerebellar hemisphere (P ataxia telangiectasia patients along with white matter tract degeneration projecting from the cerebellum into corticomotor regions. The lack of cortical involvement may reflect early-stage white matter motor pathway degeneration within young patients. © 2014 International Parkinson and Movement Disorder Society.

  4. Clinical spectrum of ataxia-telangiectasia in adulthood

    NARCIS (Netherlands)

    Verhagen, M. M. M.; Abdo, W. F.; Willemsen, M. A. A. P.; Hogervorst, F. B. L.; Smeets, D. F. C. M.; Hiel, J. A. P.; Brunt, E. R.; van Rijn, M. A.; Krakauer, D. Majoor; Oldenburg, R. A.; Broeks, A.; Last, J. I.; van't Veer, L. J.; Tijssen, M. A. J.; Dubois, A. M. I.; Kremer, H. P. H.; Weemaes, C. M. R.; Taylor, A. M. R.; van Deuren, M.

    2009-01-01

    Objective: To describe the phenotype of adult patients with variant and classic ataxia-telangiectasia (A-T), to raise the degree of clinical suspicion for the diagnosis variant A-T, and to assess a genotype-phenotype relationship for mutations in the ATM gene. Methods: Retrospective analysis of the

  5. Clinical spectrum of ataxia-telangiectasia in adulthood.

    NARCIS (Netherlands)

    Verhagen, M.M.; Abdo, W.; Willemsen, M.A.A.P.; Hogervorst, F.B.L.; Smeets, D.F.C.M.; Hiel, J.A.P.; Brunt, E.R.; Rijn, M.A. van; Majoor Krakauer, D.; Oldenburg, R.A.; Broeks, A.; Last, J.I.; Veer, L.J. van 't; Tijssen, M.A.; Dubois, A.M.; Kremer, H.P.H.; Weemaes, C.M.R.; Taylor, A.M.; Deuren, M. van

    2009-01-01

    OBJECTIVE: To describe the phenotype of adult patients with variant and classic ataxia-telangiectasia (A-T), to raise the degree of clinical suspicion for the diagnosis variant A-T, and to assess a genotype-phenotype relationship for mutations in the ATM gene. METHODS: Retrospective analysis of the

  6. Disorders of Upper Limb Movements in Ataxia-Telangiectasia.

    Directory of Open Access Journals (Sweden)

    Aasef G Shaikh

    Full Text Available Ataxia-telangiectasia is known for cerebellar degeneration, but clinical descriptions of abnormal tone, posture, and movements suggest involvement of the network between cerebellum and basal ganglia. We quantitatively assessed the nature of upper-limb movement disorders in ataxia-telangiectasia. We used a three-axis accelerometer to assess the natural history and severity of abnormal upper-limb movements in 80 ataxia-telangiectasia and 19 healthy subjects. Recordings were made during goal-directed movements of upper limb (kinetic task, while arms were outstretched (postural task, and at rest. Almost all ataxia-telangiectasia subjects (79/80 had abnormal involuntary movements, such as rhythmic oscillations (tremor, slow drifts (dystonia or athetosis, and isolated rapid movements (dystonic jerks or myoclonus. All patients with involuntary movements had both kinetic and postural tremor, while 48 (61% also had resting tremor. The tremor was present in transient episodes lasting several seconds during two-minute recording sessions of all three conditions. Percent time during which episodic tremor was present was greater for postural and kinetic tasks compared to rest. Resting tremor had higher frequency but smaller amplitude than postural and kinetic tremor. Rapid non-rhythmic movements were minimal during rest, but were triggered during sustained arm postures and goal directed arm movements suggesting they are best considered a form of dystonic jerks or action myoclonus. Advancing age did not correlate with the severity of involuntary limb movements. Abnormal upper-limb movements in ataxia-telangiectasia feature classic cerebellar impairment, but also suggest involvement of the network between the cerebellum and basal ganglia.

  7. Intrinsic mitochondrial DNA repair defects in Ataxia Telangiectasia.

    Science.gov (United States)

    Sharma, Nilesh K; Lebedeva, Maria; Thomas, Terace; Kovalenko, Olga A; Stumpf, Jeffrey D; Shadel, Gerald S; Santos, Janine H

    2014-01-01

    Ataxia Telangiectasia (A-T) is a progressive childhood disorder characterized most notably by cerebellar degeneration and predisposition to cancer. A-T is caused by mutations in the kinase ATM, a master regulator of the DNA double-strand break response. In addition to DNA-damage signaling defects, A-T cells display mitochondrial dysfunction that is thought to contribute to A-T pathogenesis. However, the molecular mechanism leading to mitochondrial dysfunction in A-T remains unclear. Here, we show that lack of ATM leads to reduced mitochondrial DNA (mtDNA) integrity and mitochondrial dysfunction, which are associated to defective mtDNA repair. While protein levels of mtDNA repair proteins are essentially normal, in the absence of ATM levels specifically of DNA ligase III (Lig3), the only DNA ligase working in mitochondria is reduced. The reduction of Lig3 is observed in different A-T patient cells, in brain and pre-B cells derived from ATM knockout mice as well as upon transient or stable knockdown of ATM. Furthermore, pharmacological inhibition of Lig3 in wild type cells phenocopies the mtDNA repair defects observed in A-T patient cells. As targeted deletion of LIG3 in the central nervous system causes debilitating ataxia in mice, reduced Lig3 protein levels and the consequent mtDNA repair defect may contribute to A-T neurodegeneration. A-T is thus the first disease characterized by diminished Lig3. Published by Elsevier B.V.

  8. Lack of mutations in the P53 gene exons 5 to 8 in ataxia-telangiectasia.

    Science.gov (United States)

    Jonveaux, P; Berger, R

    1993-04-01

    Alterations of the TP53 tumor suppressor gene are present in various human malignancies and in the dominantly inherited Li-Fraumeni syndrome. Recently, a cell cycle checkpoint pathway involving p53 and GADD45 has been identified as defective in ataxia-telangiectasia. Using single strand conformation polymorphism analysis of PCR products, we looked for TP53 mutations in DNA of patients with AT. We did not find any mutation in 6 patients, suggesting that TP53 mutations are not directly involved in the cancer susceptibility observed in AT.

  9. Neurodegeneration in ataxia-telangiectasia: Multiple roles of ATM kinase in cellular homeostasis.

    Science.gov (United States)

    Choy, Kay Rui; Watters, Dianne J

    2017-05-22

    Ataxia-telangiectasia (A-T) is characterized by neuronal degeneration, cancer, diabetes, immune deficiency, and increased sensitivity to ionizing radiation. A-T is attributed to the deficiency of the protein kinase coded by the ATM (ataxia-telangiectasia mutated) gene. ATM is a sensor of DNA double-strand breaks (DSBs) and signals to cell cycle checkpoints and the DNA repair machinery. ATM phosphorylates numerous substrates and activates many cell-signaling pathways. There has been considerable debate about whether a defective DNA damage response is causative of the neurological aspects of the disease. In proliferating cells, ATM is localized mainly in the nucleus; however, in postmitotic cells such as neurons, ATM is mostly cytoplasmic. Recent studies reveal an increasing number of roles for ATM in the cytoplasm, including activation by oxidative stress. ATM associates with organelles including mitochondria and peroxisomes, both sources of reactive oxygen species (ROS), which have been implicated in neurodegenerative diseases and aging. ATM is also associated with synaptic vesicles and has a role in regulating cellular homeostasis and autophagy. The cytoplasmic roles of ATM provide a new perspective on the neurodegenerative process in A-T. This review will examine the expanding roles of ATM in cellular homeostasis and relate these functions to the complex A-T phenotype. Developmental Dynamics, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  10. Characteristic Eye Movements in Ataxia-Telangiectasia-Like Disorder: An Explanatory Hypothesis

    Directory of Open Access Journals (Sweden)

    Pamela Federighi

    2017-11-01

    Full Text Available ObjectiveTo investigate cerebellar dysfunctions and quantitatively characterize specific oculomotor changes in ataxia-telangiectasia-like disorder (ATLD, a rare autosomal recessive disease caused by mutations in the MRE11 gene. Additionally, to further elucidate the pathophysiology of cerebellar damage in the ataxia-telangiectasia (AT spectrum disorders.MethodsSaccade dynamics, metrics, and visual fixation deficits were investigated in two Italian adult siblings with genetically confirmed ATLD. Visually guided saccades were compared with those of 40 healthy subjects. Steady fixation was tested in primary and eccentric positions. Quantitative characterization of saccade parameters, saccadic intrusions (SI, and nystagmus was performed.ResultsPatients showed abnormally hypermetric and fast horizontal saccades to the left and greater inaccuracy than healthy subjects in all saccadic eye movements. Eye movement abnormalities included slow eye movements that preceded the initial saccade. Horizontal and vertical spontaneous jerk nystagmus, gaze-evoked, and rebound nystagmus were evident. Fixation was interrupted by large square-wave jerk SI and macrosaccadic oscillations.ConclusionSlow eye movements accompanying saccades, SI, and cerebellar nystagmus are frequently seen in AT patients, additionally our ATLD patients showed the presence of fast and hypermetric saccades suggesting damage of granule cell-parallel fiber-Purkinje cell synapses of the cerebellar vermis. A dual pathogenetic mechanism involving neurodevelopmental and neurodegenerative changes is hypothesized to explain the peculiar phenotype of this disease.

  11. Human iPSC-Derived Cerebellar Neurons from a Patient with Ataxia-Telangiectasia Reveal Disrupted Gene Regulatory Networks

    Directory of Open Access Journals (Sweden)

    Sam P. Nayler

    2017-10-01

    Full Text Available Ataxia-telangiectasia (A-T is a rare genetic disorder caused by loss of function of the ataxia-telangiectasia-mutated kinase and is characterized by a predisposition to cancer, pulmonary disease, immune deficiency and progressive degeneration of the cerebellum. As animal models do not faithfully recapitulate the neurological aspects, it remains unclear whether cerebellar degeneration is a neurodevelopmental or neurodegenerative phenotype. To address the necessity for a human model, we first assessed a previously published protocol for the ability to generate cerebellar neuronal cells, finding it gave rise to a population of precursors highly enriched for markers of the early hindbrain such as EN1 and GBX2, and later more mature cerebellar markers including PTF1α, MATH1, HOXB4, ZIC3, PAX6, and TUJ1. RNA sequencing was used to classify differentiated cerebellar neurons generated from integration-free A-T and control induced pluripotent stem cells. Comparison of RNA sequencing data with datasets from the Allen Brain Atlas reveals in vitro-derived cerebellar neurons are transcriptionally similar to discrete regions of the human cerebellum, and most closely resemble the cerebellum at 22 weeks post-conception. We show that patient-derived cerebellar neurons exhibit disrupted gene regulatory networks associated with synaptic vesicle dynamics and oxidative stress, offering the first molecular insights into early cerebellar pathogenesis of ataxia-telangiectasia.

  12. Cellular and molecular response to irradiation in ataxia telangiectasia and in Fanconi`s anemia

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    Ridet, A.; Guillouf, C.; Duchaud, E.; Moustacchi, E.; Rosselli, F. [Institut Curie-Recherche, UMR 218, CNRS, 75 - Paris (France)

    1997-03-01

    Ataxia telangiectasia (AT) and Fanconi anemia (FA) are recessive genetic diseases featuring chromosomal instability, increased predisposition to cancer and in vitro hypersensitivity to ionizing radiation (AT) or DNA cross-linking agents (FA). Moreover, an in vivo hypersensitivity to {gamma}-rays exposure was reported in both syndromes. Cellular response to irradiation includes growth arrest (cell cycle modification) and cell death (by apoptosis or necrosis). Since it is generally accepted that apoptosis modulates cellular sensitivity to genotoxic stress, it was of interest to investigate the contribution of apoptosis in determining FA and AT responses to DNA Damaging Agents. The results support the contention that the in vivo hypersensitivity to radiation in these syndromes is not related to a higher rate of apoptotic cells but could be to a higher necrotic response triggering inflammatory reactions in the patients affected by this syndromes. (authors)

  13. Mutation of ataxia-telangiectasia mutated is associated with dysfunctional glutathione homeostasis in cerebellar astroglia.

    Science.gov (United States)

    Campbell, Andrew; Bushman, Jared; Munger, Joshua; Noble, Mark; Pröschel, Christoph; Mayer-Pröschel, Margot

    2016-02-01

    Astroglial dysfunction plays an important role in neurodegenerative diseases otherwise attributed to neuronal loss of function. Here we focus on the role of astroglia in ataxia-telangiectasia (A-T), a disease caused by mutations in the ataxia-telangiectasia mutated (ATM) gene. A hallmark of A-T pathology is progressive loss of cerebellar neurons, but the mechanisms that impact neuronal survival are unclear. We now provide a possible mechanism by which A-T astroglia affect the survival of cerebellar neurons. As astroglial functions are difficult to study in an in vivo setting, particularly in the cerebellum where these cells are intertwined with the far more numerous neurons, we conducted in vitro coculture experiments that allow for the generation and pharmacological manipulation of purified cell populations. Our analyses revealed that cerebellar astroglia isolated from Atm mutant mice show decreased expression of the cystine/glutamate exchanger subunit xCT, glutathione (GSH) reductase, and glutathione-S-transferase. We also found decreased levels of intercellular and secreted GSH in A-T astroglia. Metabolic labeling of l-cystine, the major precursor for GSH, revealed that a key component of the defect in A-T astroglia is an impaired ability to import this rate-limiting precursor for the production of GSH. This impairment resulted in suboptimal extracellular GSH supply, which in turn impaired survival of cerebellar neurons. We show that by circumventing the xCT-dependent import of L-cystine through addition of N-acetyl-L-cysteine (NAC) as an alternative cysteine source, we were able to restore GSH levels in A-T mutant astroglia providing a possible future avenue for targeted therapeutic intervention. © 2015 Wiley Periodicals, Inc.

  14. Inmunodeficiencia con ataxia telangiectasia: presentación de 4 casos

    Directory of Open Access Journals (Sweden)

    Ma. Victoria Guntiñas Zamora

    2004-03-01

    Full Text Available Se presentan 4 pacientes con síntomas y signos propios de una inmunodeficiencia con ataxia telangiectasia, tanto desde el punto de vista clínico como de los marcadores serológicos y celulares. En todos los casos la ataxia se evidenció cuando los pacientes comenzaron a caminar, las telangiectasias tuvieron una aparición más tardía y las infecciones fueron manifiestas desde edades muy tempranas. Los defectos inmunológicos fueron heterogéneos, tanto de células B como T. Todos los pacientes se encuentran actualmente bajo tratamiento inmunoestimulante a pesar de lo cual mantienen cuadros infecciosos frecuentes. La enfermedad neurológica progresa. Se recomienda el seguimiento estrecho de los casos por la posible aparición de complicaciones graves como enfermedad pulmonar crónica o neoplasias linforreticulares4 patients with symptoms and signs typical of an immunodeficiency with ataxia telangiectasia from the clinical point of view and from the point of view of the serological and cellular markers are presented. In all the cases, ataxia was confirmed when the patients began to walk. Telangiectasies had a later appearance and the infections manifested at very early ages. The immunological defects of B and T cells were heterogeneous. The patients maintain infectious pictures despite being under immunostimulating treatment. The neurological disease is in progress. It is recommended the close follow-up of the cases due to the possible emergence of severe complications, such as chronic pulmonary disease or lymphoreticular neoplasias.

  15. Brain glucose metabolism in adults with ataxia-telangiectasia and their asymptomatic relatives

    Science.gov (United States)

    Tomasi, Dardo; Wang, Gene-Jack; Studentsova, Yana; Margus, Brad; Crawford, Thomas O.

    2014-01-01

    Ataxia-telangiectasia is a recessive genetic disorder (ATM is the mutated gene) of childhood with severe motor impairments and whereas homozygotes manifest the disorder, heterozygotes are asymptomatic. Structural brain imaging and post-mortem studies in individuals with ataxia-telangiectasia have reported cerebellar atrophy; but abnormalities of motor control characteristic of extrapyramidal dysfunction suggest impairment of broader motor networks. Here, we investigated possible dysfunction in other brain areas in individuals with ataxia-telangiectasia and tested for brain changes in asymptomatic relatives to assess if heterozygocity affects brain function. We used positron emission tomography and 18F-fluorodeoxyglucose to measure brain glucose metabolism (quantified as µmol/100 g/min), which serves as a marker of brain function, in 10 adults with ataxia-telangiectasia, 19 non-affected adult relatives (12 siblings, seven parents) and 29 age-matched healthy controls. Statistical parametric mapping and region of interest analyses were used to compare individuals with ataxia-telangiectasia, asymptomatic relatives, and unrelated controls. We found that participants with ataxia-telangiectasia had lower metabolism in cerebellar hemispheres (14%, P ataxia-telangiectasia also had higher metabolism in globus pallidus (16%, P = 0.05), which correlated negatively with motor performance. Asymptomatic relatives had lower metabolism in anterior vermis (12%; P = 0.01) and hippocampus (19%; P = 0.002) than controls. Our results indicate that, in addition to the expected decrease in cerebellar metabolism, participants with ataxia-telangiectasia had widespread changes in metabolic rates including hyperactivity in globus pallidus indicative of basal ganglia involvement. Changes in basal ganglia metabolism offer potential insight into targeting strategies for therapeutic deep brain stimulation. Our finding of decreased metabolism in vermis and hippocampus of asymptomatic relatives

  16. Brain glucose metabolism in adults with ataxia-telangiectasia and their asymptomatic relatives.

    Science.gov (United States)

    Volkow, Nora D; Tomasi, Dardo; Wang, Gene-Jack; Studentsova, Yana; Margus, Brad; Crawford, Thomas O

    2014-06-01

    Ataxia-telangiectasia is a recessive genetic disorder (ATM is the mutated gene) of childhood with severe motor impairments and whereas homozygotes manifest the disorder, heterozygotes are asymptomatic. Structural brain imaging and post-mortem studies in individuals with ataxia-telangiectasia have reported cerebellar atrophy; but abnormalities of motor control characteristic of extrapyramidal dysfunction suggest impairment of broader motor networks. Here, we investigated possible dysfunction in other brain areas in individuals with ataxia-telangiectasia and tested for brain changes in asymptomatic relatives to assess if heterozygocity affects brain function. We used positron emission tomography and (18)F-fluorodeoxyglucose to measure brain glucose metabolism (quantified as µmol/100 g/min), which serves as a marker of brain function, in 10 adults with ataxia-telangiectasia, 19 non-affected adult relatives (12 siblings, seven parents) and 29 age-matched healthy controls. Statistical parametric mapping and region of interest analyses were used to compare individuals with ataxia-telangiectasia, asymptomatic relatives, and unrelated controls. We found that participants with ataxia-telangiectasia had lower metabolism in cerebellar hemispheres (14%, P brain stimulation. Our finding of decreased metabolism in vermis and hippocampus of asymptomatic relatives suggests that heterozygocity influences the function of these brain regions. Published by Oxford University Press on behalf of the Guarantors of Brain 2014. This work is written by US Government employees and is in the public domain in the US.

  17. Clinical and genetic features of ataxia-telangiectasia

    Energy Technology Data Exchange (ETDEWEB)

    Bundey, S. [Birmingham Maternity Hospital (United Kingdom). Clinical Genetics Unit

    1994-12-01

    There are several variants of ataxia-telangiectasia (A-T): classical A-T with marked radiation sensitivity; classical A-T with intermediate levels of radiation sensitivity; mild A-T with intermediate levels of radiation sensitivity; A-T without telangiectasia; A-T without oculomoto apraxia; and A-T with microcephaly. These disorders are probably caused by different allelic mutations, because affected sibs resemble the index patients, and because there is an association of certain haplo-types of 11q22-23 with specific phenotypes. The Nijmegen Breakage Syndrome, with its lack of ataxia, seems on clinical grounds to be a different disorder. Although A-T is almost always inherited as an autosomal recessive, there are some unusual features; an unexpectedly low parental consanguinity rate, an incidence in sibs that is < 0.25, and occurrence of disease in many different races and in the offspring of mixed race unions. Moreover, looking at haplotypes from 63 UK patients, there is a remarkably low incidence of homozygosity. An autosomal recessive condition that is deficient in parental consanguinity, and in homozygosity for the region around the gene, can be explained by J.H. Edwards` hypothesis that homozygosity for alleles at a neighbouring locus are lethal early in embryogenesis. Other possible mechanisms to explain the unusual genetic features are discussed. (author).

  18. Evaluation and Management of Pulmonary Disease in Ataxia-Telangiectasia

    Science.gov (United States)

    McGrath-Morrow, Sharon A.; Gower, W. Adam; Rothblum-Oviatt, Cynthia; Brody, Alan S.; Langston, Claire; Fan, Leland L.; Lefton-Greif, Maureen A.; Crawford, Thomas O.; Troche, Michelle; Sandlund, John T; Auwaerter, Paul G.; Easley, Blaine; Loughlin, Gerald M.; Carroll, John L.; Lederman, Howard M.

    2014-01-01

    Summary Ataxia-telangiectasia (A-T) is a rare autosomal recessive disorder caused by mutations in the ATM gene, resulting in faulty repair of breakages in double-stranded DNA. The clinical phenotype is complex, and is characterized by neurologic abnormalities, immunodeficiencies, susceptibility to malignancies, recurrent sinopulmonary infections, and cutaneous abnormalities. Lung disease is common in patients with A-T and often progresses with age and neurological decline. Diseases of the respiratory system cause significant morbidity and are a frequent cause of death in the A-T population. Lung disease in this population is thought to exhibit features of one or more of the following phenotypes: recurrent sinopulmonary infections with bronchiectasis, interstitial lung disease, and lung disease associated with neurological abnormalities. Here, we review available evidence and present expert opinion on the diagnosis, evaluation, and management of lung disease in A-T, as discussed in a recent multidisciplinary workshop. Although more data are emerging on this unique population, many recommendations are made based on similarities to other more well-studied diseases. Gaps in current knowledge and areas for future research in the field of pulmonary disease in A-T are also outlined. PMID:20583220

  19. Ataxia-telangiectasia: the pattern of cerebellar atrophy on MRI

    Energy Technology Data Exchange (ETDEWEB)

    Tavani, F. [Department of Radiology, University of Modena (Italy); Zimmerman, R.A.; Gatti, R.; Bingham, P. [Department of Radiology, Children' s Hospital of Philadelphia, 34th Street and Civic Center Boulevard, PA 19104, Philadelphia (United States); Berry, G.T. [Department of Endocrinology, Children' s Hospital of Philadelphia, 34th Street and Civic Center Boulevard, PA 19104, Philadelphia (United States); Sullivan, K. [Department of Immunology, Children' s Hospital of Philadelphia, 34th Street and Civic Center Boulevard, PA 19104, Philadelphia (United States)

    2003-05-01

    We describe MRI of the brain in 19 patients with ataxia-telangiectasia (AT) and correlate the appearances with the degree of neurologic deficit. We examined 10 male and nine female patients; 17 were aged between 2 and 12 years (mean 8 years) but a woman and her brother were 35 and 38 years old, and had a variant of AT. Ataxia was the first recognized sign of the disease in every patient. We detected the following patterns of cerebellar atrophy: in the youngest patient, aged 2 years, the study was normal; in the five next youngest patients 3-7 years of age, the lateral cerebellum and superior vermis showed the earliest changes of atrophy; and all but one of the other patients had moderate to marked diffuse atrophy of vermis and cerebellar hemispheres. There were 12 patients aged 9 years and above; one, who was normal, was 9 years old. The five patients who at the time of examination were unable to walk all had diffuse atrophy involving both vermis and cerebellar hemispheres. (orig.)

  20. Pulmonary function in children and young adults with ataxia telangiectasia.

    Science.gov (United States)

    McGrath-Morrow, Sharon A; Lederman, Howard M; Aherrera, Angela D; Lefton-Greif, Maureen A; Crawford, Thomas O; Ryan, Timothy; Wright, Jennifer; Collaco, Joseph M

    2014-01-01

    Pulmonary disease contributes to significant morbidity and mortality in people with ataxia telangiectasia (A-T). To determine the association between age and lung function in children and young adults with A-T and to identify factors associated with decreased lung function, pulmonary function tests were performed in 100 consecutive people with A-T. Children and adults ranging from 6 to 29 years of age and with the diagnosis of A-T were recruited, and underwent pulmonary function tests. The mean forced vital capacity % predicted (FVC %) in the population was 56.6 ± 20.0. Males and females between 6 and 10 years of age had similar pulmonary function. Older females were found to have significantly lower FVCs % than both older males (P pulmonary function testing on two or more occasions over an average of 2 years. In children and young adults with A-T, older females and people who required supplemental gamma globulin had significantly lower lung function by cross-sectional analysis. Stable lung function is possible over a 2-year period. Recognition of groups who are at higher risk for lower pulmonary function may help direct care and improve clinical outcomes in people with A-T. © 2013 Wiley Periodicals, Inc.

  1. Ataxia telangiectasia mutated-dependent regulation of topoisomerase II alpha expression and sensitivity to topoisomerase II inhibitor.

    Science.gov (United States)

    Tamaichi, Hiroyuki; Sato, Masaki; Porter, Andrew C G; Shimizu, Toshiaki; Mizutani, Shuki; Takagi, Masatoshi

    2013-02-01

    Topoisomerase II alpha (TOP2A) has a crucial role in proper chromosome condensation and segregation. Here we report the interaction of TOP2A with ataxia telangiectasia mutated (ATM) and its phosphorylation in an ATM-dependent manner after DNA damage. In vitro kinase assay and site-directed mutagenesis studies revealed that serine 1512 is the target of phosphorylation through ATM. Serine 1512 to Alanine mutation of TOP2A showed increased stability of the protein, retaining TOP2A activity at least with regard to cell survival activity. Ataxia telangiectasia-derived cell lines showed high levels of TOP2A that were associated with hypersensitivity to the TOP2 inhibitor etoposide. These findings suggest that ATM-dependent TOP2A modification is required for proper regulation of TOP2 stability and subsequently of the sensitivity to TOP2 inhibitor. In a lymphoblastoid cell line derived from a patient who developed MLL rearrangement, positive infant leukemia, defective ATM expression, and increased TOP2A expression were shown. It was intriguing that hypersensitivity to TOP2 inhibitor and susceptibility to MLL gene rearrangement were shown by low-dose etoposide exposure in this cell line. Thus, our findings have clinically important implications for the pathogenesis of infantile acute leukemia as well as treatment-associated secondary leukemia following exposure to TOP2 inhibitors. © 2012 Japanese Cancer Association.

  2. Assessment of impaired coordination between respiration and deglutition in children and young adults with ataxia telangiectasia.

    Science.gov (United States)

    Lefton-Greif, Maureen A; Perlman, Adrienne L; He, Xuming; Lederman, Howard M; Crawford, Thomas O

    2016-10-01

    This cross-sectional investigation aimed to assess the value of non-invasive measures of temporal respiratory-swallow coupling in individuals with ataxic swallowing. Twenty participants (11 males, 9 females; range 9-21y) with ataxia telangiectasia were presented with water and pudding boluses. Their 193 swallows were compared with 2200 swallows from 82 age-matched healthy controls. The two components of airway protection during swallowing that were analyzed were: direction of peri-deglutitive airflow and duration of deglutitive inhibition of respiratory airflow (DIORA). Safe expiratory patterns of peri-deglutitive airflow occurred significantly less often in participants with ataxia telangiectasia than in age-matched control participants (younger pdeglutitive airflow increased with age in participants in the comparison group (p=0.006), but not in those with ataxia telangiectasia (p=0.234). With age, mean duration of DIORA decreased in controls (p<0.001) but was unchanged in participants with ataxia telangiectasia (p=0.164). Non-invasive quantitative measures of respiratory-swallow coupling capture temporal relationships that plausibly contribute to airway compromise from dysphagia. Changes in respiratory-swallow coupling observed with advancing age in control participants were not seen in participants with ataxia telangiectasia. Measures of perturbations may herald swallowing problems prior to development of pulmonary and nutritional sequelae. © 2016 Mac Keith Press.

  3. Growth and nutrition in children with ataxia telangiectasia.

    Science.gov (United States)

    Stewart, Emma; Prayle, Andrew P; Tooke, Alison; Pasalodos, Sara; Suri, Mohnish; Bush, Andy; Bhatt, Jayesh M

    2016-12-01

    Ataxia telangiectasia (A-T) is a rare multisystem disease with high early mortality from lung disease and cancer. Nutritional failure adversely impacts outcomes in many respiratory diseases. Several factors influence nutrition in children with A-T. We hypothesised that children with A-T have progressive growth failure and that early gastrostomy tube feeding (percutaneous endoscopic gastrostomy, PEG) is a favourable management option with good nutritional outcomes. Data were collected prospectively on weight, height and body mass index (BMI) at the national paediatric A-T clinic. Adequacy and safety of oral intake was assessed. Nutritional advice was given at each multidisciplinary review. 101 children (51 girls) had 222 measurements (32 once, 32 twice, 24 thrice) between 2009 and 2016. Median (IQR) age was 9.3 (6.4 to 13.1) years. Mean (SD) weight, height and BMI Z-scores were respectively -1 (1.6), -1.2 (1.2) and -0.4 (1.4). 35/101 children had weight Z-scores below -2 on at least one occasion. Weight, height and BMI Z-scores declined over time. Decline was most obvious after 8 years of age. 14/101 (14%) children had a PEG, with longitudinal data available for 12. In a nested case control study, there was a trend for improvement in weight in those with a PEG (p=0.10). Patients with A-T decline in growth over time. There is an urgent need for new strategies, including an understanding of why growth falters. We suggest early proactive consideration of PEG from age 8 years onwards to prevent progressive growth failure. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  4. Novel compound heterozygous mutations in a child with Ataxia-Telangiectasia showing unrelated cerebellar disorders.

    Science.gov (United States)

    Piane, Maria; Molinaro, Anna; Soresina, Annarosa; Costa, Silvia; Maffeis, Marianna; Germani, Aldo; Pinelli, Lorenzo; Meschini, Roberta; Plebani, Alessandro; Chessa, Luciana; Micheli, Roberto

    2016-12-15

    We report the case of a 6-year-old female patient with Ataxia Telangiectasia, an extremely rare condition, who developed in addition a left cerebellar astrocytoma and a right cerebellar infarction, considered as two independent events. Children with AT have an increased risk of developing cancer, but only few cases of glioma are reported and, at our knowledge, no other case of unrelated cerebellar glioma and cerebellar infarction in with the same AT patient have been described. The molecular analysis of ATM (Ataxia Telangiectasia Mutated) gene showed that the patient is compound heterozygote for two previously unreported mutations: c.3291delC (p.Phe1097fs) at exon 25 and c.8198A>C (p.Gln2733Pro) at exon 58. The role of the identified ATM gene mutations in the pathogenesis of Ataxia Telangiectasia and the coexisting cerebellar disorders is discussed. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Role of ataxia-telangiectasia mutated (ATM) in porcine oocyte in vitro maturation.

    Science.gov (United States)

    Lin, Zi-Li; Kim, Nam-Hyung

    2015-06-01

    Ataxia-telangiectasia mutated (ATM) is critical for the DNA damage response, cell cycle checkpoints, and apoptosis. Significant effort has focused on elucidating the relationship between ATM and other nuclear signal transducers; however, little is known about the connection between ATM and oocyte meiotic maturation. We investigated the function of ATM in porcine oocytes. ATM was expressed at all stages of oocyte maturation and localized predominantly in the nucleus. Furthermore, the ATM-specific inhibitor KU-55933 blocked porcine oocyte maturation, reducing the percentages of oocytes that underwent germinal vesicle breakdown (GVBD) and first polar body extrusion. KU-55933 also decreased the expression of DNA damage-related genes (breast cancer 1, budding uninhibited by benzimidazoles 1, and P53) and reduced the mRNA and protein levels of AKT and other cell cycle-regulated genes that are predominantly expressed during G2/M phase, including bone morphogenetic protein 15, growth differentiation factor 9, cell division cycle protein 2, cyclinB1, and AKT. KU-55933 treatment decreased the developmental potential of blastocysts following parthenogenetic activation and increased the level of apoptosis. Together, these data suggested that ATM influenced the meiotic and cytoplasmic maturation of porcine oocytes, potentially by decreasing their sensitivity to DNA strand breaks, stimulating the AKT pathway, and/or altering the expression of other maternal genes. © 2015 International Federation for Cell Biology.

  6. Molecular and Functional Characterization of a Cohort of Spanish Patients with Ataxia-Telangiectasia.

    Science.gov (United States)

    Carranza, Diana; Vega, Ana Karina; Torres-Rusillo, Sara; Montero, Enrique; Martinez, Luis Javier; Santamaría, Manuel; Santos, Juan Luis; Molina, Ignacio J

    2017-03-01

    Ataxia-telangiectasia is a multisystemic disease with severe neurological affectation, immunodeficiency and telangiectasia. The disorder is caused by alterations in the ATM gene, whose size and complexity make molecular diagnosis difficult. We designed a target-enrichment next-generation sequencing strategy to characterize 28 patients from several regions of Spain. This approach allowed us to identify gene variants affecting function in 54 out of the 56 alleles analyzed, although the two unresolved alleles belong to brothers. We found 28 ATM gene mutations, of which 10 have not been reported. A total of 171 gene variants not affecting function were also found, of which 22 are reported to predispose to disease. Interestingly, all Roma (Spanish Gypsies) patients are homozygous for the same mutation and share the H3 ATM haplotype, which is strong evidence of a founder effect in this population. In addition, we generated a panel of 27 primary T cell lines from A-T patients, which revealed significant expression of ATM in two patients and traces of the protein in nine more. None of them retained residual ATM activity, and almost all T cell lines show increased or intermediate radiosensitivity.

  7. A single ataxia telangiectasia gene with a product similar to PI-3 kinase

    Energy Technology Data Exchange (ETDEWEB)

    Savitsky, K.; Bar-Shira, A.; Gilad, S.; Rotman, G.; Ziv, Y.; Vanagaite, L.; Smith, S.; Uziel, T.; Sfez, S.; Ashkenazi, M. [Tel Aviv Univ. (Israel)] [and others

    1995-06-23

    A gene, ATM, that is mutated in the autosomal recessive disorder ataxia telangiectasia (AT) was identified by positional cloning on chromosome 11q22-23. AT is characterized by cerebellar degeneration, immunodeficiency, chromosomal instability, cancer predisposition, radiation sensitivity, and cell cycle abnormalities. The disease is genetically heterogeneous, with four complementation groups that have been suspected to represent different genes. ATM, which has a transcript of 12 kilobases, was found to be mutated in AT patients from all complementation groups, indicating that it is probably the sole gene responsible for this disorder. A partial ATM complementary DNA clone of 5.9 kilobases encoded a putative protein that is similar to several yeast and mammalian phosphatidylinositol-3{prime} kinases that are involved in mitogenic signal transduction, meiotic recombination, and cell cycle control. The discovery of ATM should enhance understanding of AT and related syndromes and may allow the identification of AT heterozygotes, who are at increased risk of cancer. 54 refs., 5 figs., 1 tab.

  8. The role of the neuro-astro-vascular unit in the etiology of ataxia telangiectasia.

    Science.gov (United States)

    Meshulam, Leenoy; Galron, Ronit; Kanner, Sivan; De Pittà, Maurizio; Bonifazi, Paolo; Goldin, Miri; Frenkel, Dan; Ben-Jacob, Eshel; Barzilai, Ari

    2012-01-01

    The growing recognition that brain pathologies do not affect neurons only but rather are, to a large extent, pathologies of glial cells as well as of the vasculature opens to new perspectives in our understanding of genetic disorders of the CNS. To validate the role of the neuron-glial-vascular unit in the etiology of genome instability disorders, we report about cell death and morphological aspects of neuroglia networks and the associated vasculature in a mouse model of Ataxia Telangiectasia (A-T), a human genetic disorder that induces severe motor impairment. We found that A-T-mutated protein deficiency was consistent with aberrant astrocytic morphology and alterations of the vasculature, often accompanied by reactive gliosis. Interestingly similar findings could also be reported in the case of other genetic disorders. These observations bolster the notion that astrocyte-specific pathologies, hampered vascularization and astrocyte-endothelium interactions in the CNS could play a crucial role in the etiology of genome instability brain disorders and could underlie neurodegeneration.

  9. The role of the neuro-astro-vascular unit in the etiology of Ataxia Telangiectasia

    Directory of Open Access Journals (Sweden)

    Leenoy eMeshulam

    2012-09-01

    Full Text Available The growing recognition that brain pathologies do not affect neurons only but rather are, to a large extent, pathologies of glial cells as well as of the vasculature opens to new perspectives in our understanding of genetic disorders of the CNS. To validate the role of the neuron-glial-vascular unit in the etiology of genome instability disorders, we report about cell death and morphological aspects of neuro-glia networks and the associated vasculature in a mouse model of Ataxia Telangiectasia (A-T, a human genetic disorder that induces severe motor impairment. We found that AT-mutated protein deficiency was consistent with aberrant astrocytic morphology and alterations of the vasculature, often accompanied by reactive gliosis. Interestingly similar findings could also be reported in the case of other genetic disorders. These observations bolster the notion that astrocyte-specific pathologies, hampered vascularization and astrocyte-endothelium interactions in the CNS could play a crucial role in the etiology of genome instability brain disorders and could underlie neurodegeneration.

  10. Role of chromatin structure modulation by the histone deacetylase inhibitor trichostatin A on the radio-sensitivity of ataxia telangiectasia

    Energy Technology Data Exchange (ETDEWEB)

    Meschini, Roberta, E-mail: meschini@unitus.it; Morucci, Elisa; Berni, Andrea; Lopez-Martinez, Wilner; Palitti, Fabrizio

    2015-07-15

    Highlights: • Role of chromatin compaction on chromosomal instability. • Reduced radiation-induced clastogenicity in Ataxia telangiectasia cell lines. • Histone tails hyperacetylation reduces heterochromatin content favouring DSBs repair. - Abstract: At present, a lot is known about biochemical aspects of double strand breaks (DBS) repair but how chromatin structure affects this process and the sensitivity of DNA to DSB induction is still an unresolved question. Ataxia telangiectasia (A-T) patients are characterised by very high sensitivity to DSB-inducing agents such as ionising radiation. This radiosensitivity is revealed with an enhancement of chromosomal instability as a consequence of defective DNA repair for a small fraction of breaks located in the heterochromatin, where they are less accessible. Besides, recently it has been reported that Ataxia Telangiectasia Mutated (ATM) mediated signalling modifies chromatin structure. In order to study the impact of chromatin compaction on the chromosomal instability of A-T cells, the response to trichostatin-A, an histone deacetylase inhibitor, in normal and A-T lymphoblastoid cell lines was investigated testing its effect on chromosomal aberrations, cell cycle progression, DNA damage and repair after exposure to X-rays. The results suggest that the response to both trichostatin-A pre- and continuous treatments is independent of the presence of either functional or mutated ATM protein, as the reduction of chromosomal damage was found also in the wild-type cell line. The presence of trichostatin-A before exposure to X-rays could give rise to prompt DNA repair functioning on chromatin structure already in an open conformation. Differently, trichostatin-A post-treatment causing hyperacetylation of histone tails and reducing the heterochromatic DNA content might diminish the requirement for ATM and favour DSBs repair reducing chromosomal damage only in A-T cells. This fact could suggest that trichostatin-A post

  11. Antioxidant vitamins intake, ataxia telangiectasia mutated (ATM) genetic polymorphisms, and breast cancer risk.

    Science.gov (United States)

    Lee, Sang-Ah; Lee, Kyoung-Mu; Lee, Seung-Joon; Yoo, Keun-Young; Park, Sue Kyung; Noh, Dong-Young; Ahn, Sei-Hyun; Kang, Daehee

    2010-01-01

    Ataxia telangiectasia mutated (ATM) cells exist under a constant state of oxidative stress with high levels of reactive oxygen species, which are removed by cellular antioxidant vitamins. We investigated the independent and combined effect of antioxidant vitamins intake and the ATM genotype or diplotype on the breast cancer risk. Analyses included 323 cases and age-matched controls who participated in the Korean Breast Cancer Study during 2001-2003 with complete dietary information. The vitamin A (P vitamins including vitamin B(2) (P = 0.01), vitamin C (P intake. No five single nucleotide polymorphisms (ATM-5144A > T (rs228589), IVS21 + 1049T > C (rs664677), IVS33-55T > C (rs664982), IVS34+60G > A (rs664143), and 3393T > G (rs4585)) studied showed significant differences in their allele frequencies between the cases and controls. On the other hand, compared with the diploid of ATTGT/ATTGT, as the number of ATTGT haplotype decreased, the risk of breast cancer increased (P = 0.04). The association between ATM diplotype and the breast cancer risk was predominantly among women with low intake of antioxidant vitamins including vitamin A, vitamin C, and folic acid. This study suggested that some antioxidant vitamins intake may modify the effect of ATM diplotype on the breast cancer risk among Korean women.

  12. A novel porcine model of ataxia telangiectasia reproduces neurological features and motor deficits of human disease.

    Science.gov (United States)

    Beraldi, Rosanna; Chan, Chun-Hung; Rogers, Christopher S; Kovács, Attila D; Meyerholz, David K; Trantzas, Constantin; Lambertz, Allyn M; Darbro, Benjamin W; Weber, Krystal L; White, Katherine A M; Rheeden, Richard V; Kruer, Michael C; Dacken, Brian A; Wang, Xiao-Jun; Davis, Bryan T; Rohret, Judy A; Struzynski, Jason T; Rohret, Frank A; Weimer, Jill M; Pearce, David A

    2015-11-15

    Ataxia telangiectasia (AT) is a progressive multisystem disorder caused by mutations in the AT-mutated (ATM) gene. AT is a neurodegenerative disease primarily characterized by cerebellar degeneration in children leading to motor impairment. The disease progresses with other clinical manifestations including oculocutaneous telangiectasia, immune disorders, increased susceptibly to cancer and respiratory infections. Although genetic investigations and physiological models have established the linkage of ATM with AT onset, the mechanisms linking ATM to neurodegeneration remain undetermined, hindering therapeutic development. Several murine models of AT have been successfully generated showing some of the clinical manifestations of the disease, however they do not fully recapitulate the hallmark neurological phenotype, thus highlighting the need for a more suitable animal model. We engineered a novel porcine model of AT to better phenocopy the disease and bridge the gap between human and current animal models. The initial characterization of AT pigs revealed early cerebellar lesions including loss of Purkinje cells (PCs) and altered cytoarchitecture suggesting a developmental etiology for AT and could advocate for early therapies for AT patients. In addition, similar to patients, AT pigs show growth retardation and develop motor deficit phenotypes. By using the porcine system to model human AT, we established the first animal model showing PC loss and motor features of the human disease. The novel AT pig provides new opportunities to unmask functions and roles of ATM in AT disease and in physiological conditions. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  13. A high frequency of distinct ATM gene mutations in ataxia-telangiectasia

    Energy Technology Data Exchange (ETDEWEB)

    Wright, J.; Teraoka, S.; Concannon, P. [Univ. of Washington School of Medicine, Seattle, WA (United States)] [and others

    1996-10-01

    The clinical features of the autosomal recessive disorder ataxia-telangiectasia (AT) include a progressive cerebellar ataxia, hypersensitivity to ionizing radiation, and an increased susceptibility to malignancies. Epidemiological studies have suggested that AT heterozygotes may also be at increased risk for malignancy, possibly as a consequence of radiation exposure. A gene mutated in AT patients (ATM) has recently been isolated, making mutation screening in both patients and the general population possible. Because of the relatively large size of the ATM gene, the design of screening programs will depend on the types and distribution of mutations in the general population. In this report, we describe 30 mutations identified in a panel of unrelated AT patients and controls. Twenty-five of the 30 were distinct, and most patients were compound heterozygotes. The most frequently detected mutation was found in three different families and had previously been reported in five others. This corresponds to a frequency of 8% of all reported ATM mutations. Twenty-two of the alterations observed would be predicted to lead to protein truncation at sites scattered throughout the molecule. Two fibroblast cell lines, which displayed normal responses to ionizing radiation, also proved to be heterozygous for truncation mutations of ATM. These observations suggest that the carrier frequency of ATM mutations may be sufficiently high to make population screening practical. However, such screening may need to be done prospectively, that is, by searching for new mutations rather than by screening for just those already identified in AT families. 33 refs., 1 fig., 1 tab.

  14. Linear growth and endocrine function in children with ataxia telangiectasia

    Directory of Open Access Journals (Sweden)

    Mohammad Ehlayel

    2014-01-01

    Full Text Available Introduction: Ataxia telangiectasia (AT is a rare, genetic, primary immune deficiency disease characterized by immunodeficiency and neurological manifestations, with an increased tendency to infection, malignancy, and autoimmune diseases. Both growth delay and endocrine abnormalities are occasionally reported in these patients. Patients and Methods: We studied growth parameters height (Ht, weight, body mass index (BMI and calculated the Ht standard deviation scores (HtSDS of 13 patients (age 7.7 ± 3.5 years-age range: 3-14.5 years with AT in relation to their mid-parental Ht SDS (MPHtSDS. We measured their serum calcium (Ca, phosphorus (PO4, alkaline phosphatase, alanine transferase (ALT, serum ferritin, creatinine and albumin concentrations. Endocrine investigations included the assessment of serum free thyroxine (FT4, thyrotropin (TSH, insulin-like growth factor-I (IGF-I and morning cortisol. Complete blood count and serum immunoglobulins (IgG, IgM and IgA antibodies were also measured. Growth data were correlated to hormonal and immune data. Results: About 31% of patients with AT had short stature (HtSDS <−2. However, their MPHtSDS denoted that their short stature was familial because four out of 13 had MPHtSDS <−2. They had low BMI, and two of them had low serum albumin and IGF-I, denoting malnutrition or disturbed growth hormone secretion. Elevated serum ALT and ferritin in some patients suggest immune-related inflammation in the liver. 30% of patients had high TSH, two of them had low FT4 diagnosing overt (15% and sub-clinical (15% hypothyroidism. Anti-thyroid peroxidase antibodies were high in two out of 13 patients denoting immune-related thyroid aggression. Eight out of 13 patients had Vitamin D deficiency (<20 ng/ml however, their serum Ca and PO 4 levels were in the normal range. One adolescent girl (14.5 years had hyper-gonadotropic hypogonadism (low estradiol and high follicle stimulating hormone. All patients had normal 8 AM

  15. NAD(+) Replenishment Improves Lifespan and Healthspan in Ataxia Telangiectasia Models via Mitophagy and DNA Repair

    DEFF Research Database (Denmark)

    Fang, Evandro Fei; Kassahun, Henok; Croteau, Deborah L

    2016-01-01

    Ataxia telangiectasia (A-T) is a rare autosomal recessive disease characterized by progressive neurodegeneration and cerebellar ataxia. A-T is causally linked to defects in ATM, a master regulator of the response to and repair of DNA double-strand breaks. The molecular basis of cerebellar atrophy...

  16. Cognitive and speech-language performance in children with ataxia telangiectasia

    NARCIS (Netherlands)

    Vinck, A.; Verhagen, M.M.; Gerven, M.; Groot, I.J.M. de; Weemaes, C.M.R.; Maassen, B.A.M.; Willemsen, M.A.A.P.

    2011-01-01

    OBJECTIVE: To describe cognitive and speech-language functioning of patients with ataxia-telangiectasia (A-T) in relation to their deteriorating (oculo)motor function. DESIGN: Observational case series. METHODS: Cognitive functioning, language, speech and oral-motor functioning were examined in

  17. Cognitive and speech-language performance in children with ataxia telangiectasia

    NARCIS (Netherlands)

    Vinck, Anja; Verhagen, Mijke M. M.; van Gerven, Marjo; de Groot, Imelda J. M.; Weemaes, Corry M. R.; Maassen, Ben A. M.; Willemsen, Michel A. A. P.

    2011-01-01

    Objective: To describe cognitive and speech-language functioning of patients with ataxia-telangiectasia (A-T) in relation to their deteriorating (oculo)motor function. Design: Observational case series. Methods: Cognitive functioning, language, speech and oral-motor functioning were examined in

  18. Motor pathway degeneration in young ataxia telangiectasia patients: A diffusion tractography study

    Directory of Open Access Journals (Sweden)

    Ishani Sahama

    2015-01-01

    Conclusions: Whole tract analysis of the corticomotor, corticospinal and somatosensory pathways in ataxia telangiectasia showed significant white matter degeneration along the entire length of motor circuits, highlighting that ataxia–telangiectasia gene mutation impacts the cerebellum and multiple other motor circuits in young patients.

  19. Oxidative stress-driven pulmonary inflammation and fibrosis in a mouse model of human ataxia-telangiectasia

    Directory of Open Access Journals (Sweden)

    Ruth Duecker

    2018-04-01

    Full Text Available Lung failure is responsible for significant morbidity and is a frequent cause of death in ataxia-telangiectasia (A-T. Disturbance in the redox balance of alveolar epithelial cells must be considered as a causal factor for respiratory disease in A-T. To investigate bronchoalveolar sensitivity to reactive oxygen species (ROS and ROS-induced DNA damage, we used bleomycin (BLM to induce experimental inflammation and fibrotic changes in the Atm-deficient mouse model.BLM or saline was administered by oropharyngeal instillation into the lung of Atm-deficient mice and wild-type mice. Mice underwent pulmonary function testing at days 0, 9, and 28, and bronchoalveolar lavage (BAL was analysed for cell distribution and cytokines. Lung tissue was analysed by histochemistry.BLM administration resulted in a tremendous increase in lung inflammation and fibrotic changes in the lung tissue of Atm-deficient mice and was accompanied by irreversible deterioration of lung function. ATM (ataxia telangiectasia mutated deficiency resulted in reduced cell viability, a delay in the resolution of γH2AX expression and a significant increase in intracellular ROS in pulmonary epithelial cells after BLM treatment. This was confirmed in the human epithelial cell line A549 treated with the ATM-kinase inhibitor KU55933.Our results demonstrate high bronchoalveolar sensitivity to ROS and ROS-induced DNA damage in the Atm-deficient mouse model and support the hypothesis that ATM plays a pivotal role in the control of oxidative stress-driven lung inflammation and fibrosis. Keywords: Pulmonary inflammation, Lung fibrosis, Mice, Oxidative stress

  20. Ataxia telangiectasia mutated (ATM) interacts with p400 ATPase for an efficient DNA damage response.

    Science.gov (United States)

    Smith, Rebecca J; Savoian, Matthew S; Weber, Lauren E; Park, Jeong Hyeon

    2016-11-04

    Ataxia telangiectasia mutated (ATM) and TRRAP proteins belong to the phosphatidylinositol 3-kinase-related kinase family and are involved in DNA damage repair and chromatin remodeling. ATM is a checkpoint kinase that is recruited to sites of DNA double-strand breaks where it phosphorylates a diverse range of proteins that are part of the chromatin and DNA repair machinery. As an integral subunit of the TRRAP-TIP60 complexes, p400 ATPase is a chromatin remodeler that is also targeted to DNA double-strand break sites. While it is understood that DNA binding transcriptional activators recruit p400 ATPase into a regulatory region of the promoter, how p400 recognises and moves to DNA double-strand break sites is far less clear. Here we investigate a possibility whether ATM serves as a shuttle to deliver p400 to break sites. Our data indicate that p400 co-immunoprecipitates with ATM independently of DNA damage state and that the N-terminal domain of p400 is vital for this interaction. Heterologous expression studies using Sf9 cells revealed that the ATM-p400 complex can be reconstituted without other mammalian bridging proteins. Overexpression of ATM-interacting p400 regions in U2OS cells induced dominant negative effects including the inhibition of both DNA damage repair and cell proliferation. Consistent with the dominant negative effect, the stable expression of an N-terminal p400 fragment showed a decrease in the association of p400 with ATM, but did not alter the association of p400 with TRRAP. Taken together, our findings suggest that a protein-protein interaction between ATM and p400 ATPase occurs independently of DNA damage and contributes to efficient DNA damage response and repair.

  1. Germline Mutations of the Ataxia-Telangiectasia Gene, ATM, as a Risk Factor for Radiation-Associated Breast Cancer

    National Research Council Canada - National Science Library

    Offit, Kenneth

    1998-01-01

    This project is a case-control study designed to determine whether or not the presence of a germline mutation in ATM, the gene responsible for ataxia-telangiectasia, significantly increases the risk...

  2. Motor pathway degeneration in young ataxia telangiectasia patients: A diffusion tractography study.

    Science.gov (United States)

    Sahama, Ishani; Sinclair, Kate; Fiori, Simona; Doecke, James; Pannek, Kerstin; Reid, Lee; Lavin, Martin; Rose, Stephen

    2015-01-01

    Our understanding of the effect of ataxia-telangiectasia mutated gene mutations on brain structure and function is limited. In this study, white matter motor pathway integrity was investigated in ataxia telangiectasia patients using diffusion MRI and probabilistic tractography. Diffusion MRI were obtained from 12 patients (age range: 7-22 years, mean: 12 years) and 12 typically developing age matched participants (age range 8-23 years, mean: 13 years). White matter fiber tracking and whole tract statistical analyses were used to assess quantitative fractional anisotropy and mean diffusivity differences along the cortico-ponto-cerebellar, cerebellar-thalamo-cortical, somatosensory and lateral corticospinal tract length in patients using a linear mixed effects model. White matter tract streamline number and apparent fiber density in patient and control tracts were also assessed. Reduced fractional anisotropy along all analyzed patient tracts were observed (p < 0.001). Mean diffusivity was significantly elevated in anterior tract locations but was reduced within cerebellar peduncle regions of all patient tracts (p < 0.001). Reduced tract streamline number and tract volume in the left and right corticospinal and somatosensory tracts were observed in patients (p < 0.006). In addition, reduced apparent fiber density in the left and right corticospinal and right somatosensory tracts (p < 0.006) occurred in patients. Whole tract analysis of the corticomotor, corticospinal and somatosensory pathways in ataxia telangiectasia showed significant white matter degeneration along the entire length of motor circuits, highlighting that ataxia-telangiectasia gene mutation impacts the cerebellum and multiple other motor circuits in young patients.

  3. Assessment of impaired coordination between respiration and deglutition in children and young adults with ataxia telangiectasia

    Science.gov (United States)

    Lefton-Greif, Maureen A; Perlman, Adrienne L; He, Xuming; Lederman, Howard M; Crawford, Thomas O

    2016-01-01

    AIM This cross-sectional investigation aimed to assess the value of non-invasive measures of temporal respiratory–swallow coupling in individuals with ataxic swallowing. METHOD Twenty participants with ataxia telangiectasia were presented with water and pudding boluses. Their 193 swallows were compared with 2200 swallows from 82 age-matched healthy controls. The two components of airway protection during swallowing that were analyzed were: direction of peri-deglutitive airflow and duration of deglutitive inhibition of respiratory airflow (DIORA). RESULTS Safe expiratory patterns of peri-deglutitive airflow occurred significantly less often in participants with ataxia telangiectasia than in age-matched control participants (younger pataxia telangiectasia (p=0.234). With age, mean duration of DIORA decreased in controls (pataxia telangiectasia (p=0.164). INTERPRETATION Non-invasive quantitative measures of respiratory–swallow coupling capture temporal relationships that plausibly contribute to airway compromise from dysphagia. Changes in respiratory–swallow coupling observed with advancing age in control participants were not seen in participants with ataxia telangiectasia. Measures of perturbations may herald swallowing problems prior to development of pulmonary and nutritional sequelae. PMID:27214374

  4. Prenatal diagnosis of ataxia-telangiectasia and Nijmegen Breakage Syndrome by the assay of radioresistant DNA synthesis

    Energy Technology Data Exchange (ETDEWEB)

    Kleijer, W.J.; Kraan, M. van der; Los, F.J. [Erasmus Univ., Rotterdam (Netherlands). Dept. of Clinical Genetics; Jaspers, N.G.J. [Erasmus Univ., Rotterdam (Netherlands). Lab. of Cell Biology and Genetics

    1994-12-01

    Prenatal diagnosis was performed in 16 pregnancies at risk of ataxia-telangiectasia (A-T) or Nijmegen Breakage Syndrome (NBS). Radioresistant DNA synthesis (RDS) was investigated in cultured chorionic villus (CV) cells and/or amniotic fluid (AF) cells. In four pregnancies, an affected foetus was diagnosed with increased RDS in cultured CV cells. In three of the four cases confirmation of the diagnosis was obtained by analysis of AF cells and/or skin fibroblasts from the foetus cultured after termination of the pregnancy; in the fourth case a fibroblast culture from the aborted foetus failed. In one case, only AF cells could be analysed in a late stage of pregnancy; pregnancy was terminated due to intermediate/equivocal results but the foetus fibroblasts showed normal RDS. Normal RDS was demonstrated in the other 11 pregnancies at 25% risk either by analysis of CB cells (nine cases) or of AF cells (two cases). In some cases the (normal) results on the CV cells were corroborated by subsequent analysis of Af cells. The results suggest that RDS analysis of CV cells allows reliable prenatal diagnosis of A-T/NBS. However, amniocentesis may be necessary to confirm normal results on CV cells if the foetus is female (because of the risk of maternal cell contamination) or in the rare case of equivocal results. (author).

  5. Gastric outlet obstruction due to adenocarcinoma in a patient with Ataxia-Telangiectasia syndrome: a case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Hammond Sue

    2009-03-01

    Full Text Available Abstract Background Ataxia-Telangiectasia syndrome is characterized by progressive cerebellar dysfunction, conjuctival and cutaneous telangiectasias, severe immune deficiencies, premature aging and predisposition to cancer. Clinical and radiographic evaluation for malignancy in ataxia-telangiectasia patients is usually atypical, leading to delays in diagnosis. Case presentation We report the case of a 20 year old ataxia-telangiectasia patient with gastric adenocarcinoma that presented as complete gastric outlet obstruction. Conclusion A literature search of adenocarcinoma associated with ataxia-telangiectasia revealed 6 cases. All patients presented with non-specific gastrointestinal complaints suggestive of ulcer disease. Although there was no correlation between immunoglobulin levels and development of gastric adenocarcinoma, the presence of chronic gastritis and intestinal metaplasia seem to lead to the development of gastric adenocarcinoma. One should consider adenocarcinoma in any patient with ataxia-telangiectasia who presents with non-specific gastrointestinal complaints, since this can lead to earlier diagnosis.

  6. Dano e reparo de dna em indivíduos com ataxia-telangiectasia e em seus pais heterozigotos

    OpenAIRE

    Roberta Passos Palazzo

    2010-01-01

    A presente pesquisa pretende demonstrar evidências capazes de contribuir para o entendimento dos mecanismos envolvidos na ataxia-telangiectasia, bem como oferecer dados que auxiliem na implementação de técnicas complementares ao diagnóstico desta síndrome. Desde a descoberta do gene envolvido na ataxia-telangiectasia (o gene ATM), muito conhecimento tem sido acumulado, especialmente sobre os mecanismos moleculares envolvidos na síndrome, bem como nas respectivas doenças relacionadas. Este est...

  7. Ataxia-telangiectasia mutated and the Mre11-Rad50-NBS1 complex: promising targets for radiosensitization.

    Science.gov (United States)

    Kuroda, Shinji; Urata, Yasuo; Fujiwara, Toshiyoshi

    2012-01-01

    Radiotherapy plays a central part in cancer treatment, and use of radiosensitizing agents can greatly enhance this modality. Although studies have shown that several chemotherapeutic agents have the potential to increase the radiosensitivity of tumor cells, investigators have also studied a number of molecularly targeted agents as radiosensitizers in clinical trials based on reasonably promising preclinical data. Recent intense research into the DNA damage-signaling pathway revealed that ataxia-telangiectasia mutated (ATM) and the Mre11-Rad50-NBS1 (MRN) complex play central roles in DNA repair and cell cycle checkpoints and that these molecules are promising targets for radiosensitization. Researchers recently developed three ATM inhibitors (KU-55933, CGK733, and CP466722) and an MRN complex inhibitor (mirin) and showed that they have great potential as radiosensitizers of tumors in preclinical studies. Additionally, we showed that a telomerase-dependent oncolytic adenovirus that we developed (OBP-301 [telomelysin]) produces profound radiosensitizing effects by inhibiting the MRN complex via the adenoviral E1B55kDa protein. A recent Phase I trial in the United States determined that telomelysin was safe and well tolerated in humans, and this agent is about to be tested in combination with radiotherapy in a clinical trial based on intriguing preclinical data demonstrating that telomelysin and ionizing radiation can potentiate each other. In this review, we highlight the great potential of ATM and MRN complex inhibitors, including telomelysin, as radiosensitizing agents.

  8. Nutritional status of patients with ataxia-telangiectasia: A case for early and ongoing nutrition support and intervention.

    Science.gov (United States)

    Ross, Lynda J; Capra, Sandra; Baguley, Brenton; Sinclair, Kate; Munro, Kate; Lewindon, Peter; Lavin, Martin

    2015-08-01

    Ataxia-telangiectasia (A-T) is a rare genomic syndrome resulting in severe disability. Chronic childhood disorders can profoundly influence growth and development. Nutrition-related issues in A-T are not well described, and there are no nutritional guidelines. This study investigated the nutrition-related characteristics and behaviours of Australian A-T patients attending a national clinic. A cross-sectional analysis of 13 A-T patients (nine females; aged: 4-23 years): nutritional status was assessed by anthropometric and body cell mass (BCM) calculations. Parents reported their child's diet history and physical and behavioural factors that affect nutrition including fatigue and need for assistance. Ten (77%) had short stature (height for age z scores underweight for height (weight/height z scores nutritional barriers as chronic tiredness and the need for care giver assistance with meals. This study confirms profound malnutrition in Australian A-T patients. Poor intakes and diet quality suggest the need for early nutrition intervention. Ongoing support for families and early discussions on tube feeding are required to address changing needs in childhood and likely nutritional decline into adulthood. A prospective study is required to assess feasibility and effectiveness of nutrition interventions in young people with A-T. © 2015 The Authors. Journal of Paediatrics and Child Health © 2015 Paediatrics and Child Health Division (Royal Australasian College of Physicians).

  9. Risk for Sporadic Breast Cancer in Ataxia Telangiectasia Heterozygotes

    Science.gov (United States)

    2001-08-01

    6 7 gas 9 a poly p73a poly p73e poly p73N FiG. 1. Endogenous p73 a and P proteins are detectable by various p73-specific antibodies. Immunoblots of...even p73-specific targets. One example may be the Aquaporin 3 gene, a glycerol and solute transporter, which is greatly preferred by the p7313...model in which some DNA damage signals are channeled through c-abl to p73. Hence, one would predict that p73-deficient cells should have defective DNA

  10. Lymphoid tumours and breast cancer in ataxia telangiectasia; substantial protective effect of residual ATM kinase activity against childhood tumours

    NARCIS (Netherlands)

    Reiman, A.; Srinivasan, V.; Barone, G.; Last, J.I.; Wootton, L.L.; Davies, E.G.; Verhagen, M.M.; Willemsen, M.A.A.P.; Weemaes, C.M.R.; Byrd, P.J.; Izatt, L.; Easton, D.F.; Thompson, D.J.; Taylor, A.M.

    2011-01-01

    BACKGROUND: Immunodeficiency in ataxia telangiectasia (A-T) is less severe in patients expressing some mutant or normal ATM kinase activity. We, therefore, determined whether expression of residual ATM kinase activity also protected against tumour development in A-T. METHODS: From a total of 296

  11. Ataxia Telangiectasia

    Science.gov (United States)

    ... Research Funded by NINDS Basic Neuroscience Clinical Research Translational Research Research at NINDS Focus on Research Alzheimer's & Related Dementias Bioengineering Epilepsy Health Disparities Neural Interfaces Parkinson's Disease Spinal Cord Injury ...

  12. Ataxia Telangiectasia

    Science.gov (United States)

    ... Español 1-800-4-CANCER Live Chat Publications Dictionary Menu Contact Dictionary Search About Cancer Causes and Prevention Risk Factors ... my chances of getting breast cancer? What about environmental sources of radiation, such as cellular phones? Should ...

  13. Bladder Wall Telangiectasia in a Patient with Ataxia-Telangiectasia and How to Manage?

    Directory of Open Access Journals (Sweden)

    Fatma Deniz Aygün

    2015-01-01

    Full Text Available Ataxia-telangiectasia (A-T is a rare neurodegenerative, inherited disease causing severe morbidity. Oculocutaneous telangiectasias are almost constant findings among the affected cases as telangiectasia is considered the main clinical finding for diagnosis. Vascular abnormalities in organs have been reported infrequently but bladder wall telangiectasias are extremely rare. We aimed to report recurrent hemorrhage from bladder wall telangiectasia in a 9-year-old boy with A-T who had received intravenous cyclophosphamide for non-Hodgkin’s lymphoma. Since A-T patients are known to be more susceptible to chemical agents, we suggested that possibly cyclophosphamide was the drug which induced bladder wall injury in this patient.

  14. A haplotype common to intermediate radiosensitivity variants of ataxia-telangiectasia in the UK

    Energy Technology Data Exchange (ETDEWEB)

    Taylor, A.M.R.; McConville, C.M.; Byrd, P.J. [Birmingham Univ. (United Kingdom). Medical School; Rotman, G.; Shiloh, Y. [Tel Aviv Univ. (Israel). Sackler School of Medicine

    1994-12-01

    In a study of ataxia-telangiectasia (A-T) in the UK, patients in10 out of 60 families were shown to have a much lower level of chromosomal radiosensitivity compared with the majority of patients. In some patients the level of radiosensitivity was hardly distinguishable from normal. Patients in this group, however, could be distinguished clinically from the majority either by the later onset of severe cerebellar features or the slower rate of progress of the disorder. By using highly polymorphic microsatellite repeat markers a chromosome 11q22-23 haplotype common to the majority of these patients, and not occurring in any non-A-T chromosome in 60 families, was identified on one chromosome. The haplotype probably defines the region of the A-T gene in these families and the mutation associated with this haplotype may be much less severe than the second mutation thereby producing the slightly milder phenotype. (author).

  15. Oxidative stress, mitochondrial abnormalities and antioxidant defense in Ataxia-telangiectasia, Bloom syndrome and Nijmegen breakage syndrome.

    Science.gov (United States)

    Maciejczyk, Mateusz; Mikoluc, Bozena; Pietrucha, Barbara; Heropolitanska-Pliszka, Edyta; Pac, Malgorzata; Motkowski, Radosław; Car, Halina

    2017-04-01

    Rare pleiotropic genetic disorders, Ataxia-telangiectasia (A-T), Bloom syndrome (BS) and Nijmegen breakage syndrome (NBS) are characterised by immunodeficiency, extreme radiosensitivity, higher cancer susceptibility, premature aging, neurodegeneration and insulin resistance. Some of these functional abnormalities can be explained by aberrant DNA damage response and chromosomal instability. It has been suggested that one possible common denominator of these conditions could be chronic oxidative stress caused by endogenous ROS overproduction and impairment of mitochondrial homeostasis. Recent studies indicate new, alternative sources of oxidative stress in A-T, BS and NBS cells, including NADPH oxidase 4 (NOX4), oxidised low-density lipoprotein (ox-LDL) or Poly (ADP-ribose) polymerases (PARP). Mitochondrial abnormalities such as changes in the ultrastructure and function of mitochondria, excess mROS production as well as mitochondrial damage have also been reported in A-T, BS and NBS cells. A-T, BS and NBS cells are inextricably linked to high levels of reactive oxygen species (ROS), and thereby, chronic oxidative stress may be a major phenotypic hallmark in these diseases. Due to the presence of mitochondrial disturbances, A-T, BS and NBS may be considered mitochondrial diseases. Excess activity of antioxidant enzymes and an insufficient amount of low molecular weight antioxidants indicate new pharmacological strategies for patients suffering from the aforementioned diseases. However, at the current stage of research we are unable to ascertain if antioxidants and free radical scavengers can improve the condition or prolong the survival time of A-T, BS and NBS patients. Therefore, it is necessary to conduct experimental studies in a human model. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Oxidative stress, mitochondrial abnormalities and antioxidant defense in Ataxia-telangiectasia, Bloom syndrome and Nijmegen breakage syndrome

    Directory of Open Access Journals (Sweden)

    Mateusz Maciejczyk

    2017-04-01

    Full Text Available Rare pleiotropic genetic disorders, Ataxia-telangiectasia (A-T, Bloom syndrome (BS and Nijmegen breakage syndrome (NBS are characterised by immunodeficiency, extreme radiosensitivity, higher cancer susceptibility, premature aging, neurodegeneration and insulin resistance. Some of these functional abnormalities can be explained by aberrant DNA damage response and chromosomal instability. It has been suggested that one possible common denominator of these conditions could be chronic oxidative stress caused by endogenous ROS overproduction and impairment of mitochondrial homeostasis. Recent studies indicate new, alternative sources of oxidative stress in A-T, BS and NBS cells, including NADPH oxidase 4 (NOX4, oxidised low-density lipoprotein (ox-LDL or Poly (ADP-ribose polymerases (PARP. Mitochondrial abnormalities such as changes in the ultrastructure and function of mitochondria, excess mROS production as well as mitochondrial damage have also been reported in A-T, BS and NBS cells. A-T, BS and NBS cells are inextricably linked to high levels of reactive oxygen species (ROS, and thereby, chronic oxidative stress may be a major phenotypic hallmark in these diseases. Due to the presence of mitochondrial disturbances, A-T, BS and NBS may be considered mitochondrial diseases. Excess activity of antioxidant enzymes and an insufficient amount of low molecular weight antioxidants indicate new pharmacological strategies for patients suffering from the aforementioned diseases. However, at the current stage of research we are unable to ascertain if antioxidants and free radical scavengers can improve the condition or prolong the survival time of A-T, BS and NBS patients. Therefore, it is necessary to conduct experimental studies in a human model.

  17. Genetic, physical and functional analysis of the ataxia-telangiectasia locus on chromosome 11q22-23

    Energy Technology Data Exchange (ETDEWEB)

    Shiloh, Y.; Ziv, Y.; Savitski, K. [Tel Aviv Univ. (Israel)] [and others

    1994-09-01

    Ataxia-telangiectasia (A-T) is an autosomal recessive multisystem disorder featuring cerebellar degeneration, immunodeficiency, chromosomal instability, cancer susceptibility, and radiosensitivity. Four complementation groups have been observed in A-T. The two major groups, A and C, were localized to chromosome 11q22-23, and the other two, D and E, may map to the same chromosomal region. We developed an integrated system of positional and complementation cloning to identify the A-T gene(s). The A-T region was saturated with microsatellite markers by physically mapping markers generated at random by other labs and by identifying new polymorphic CA-repeats in YAC clones obtained from this region. According to recent linkage data based on these markers and linkage disequilibrium analysis in Moroccan Jewish A-T patients, the A-T(A) and A-T(C) mutations are contained within a 2 Mb interval between D11S1819 and D11S1960. This interval was cloned in YAC and cosmid contigs, and transcribed sequences were identified using the following methods: screening of cDNA libraries using cosmid clones; magnetic bead capture using YAC and cosmid clones; direct selection of cDNA clones using YAC clones immobilized on a solid matrix; and 3{prime} exon trapping. Preliminary results indicate that the A-T region is rich in transcribed sequences. Structural and functional analysis of these genes is being carried out by sequence analysis, by physical mapping using the cosmid contigs, and by testing their ability to complement the radiomimetic sensitivity of A-T cells.

  18. Cognitive and speech-language performance in children with ataxia telangiectasia.

    Science.gov (United States)

    Vinck, Anja; Verhagen, Mijke M M; Gerven, Marjo van; de Groot, Imelda J M; Weemaes, Corry M R; Maassen, Ben A M; Willemsen, Michel A A P

    2011-01-01

    To describe cognitive and speech-language functioning of patients with ataxia-telangiectasia (A-T) in relation to their deteriorating (oculo)motor function. Observational case series. Cognitive functioning, language, speech and oral-motor functioning were examined in eight individuals with A-T (six boys, two girls), taking into account the confounding effects of motor functioning on test performance. All patients, except the youngest one, suffered from mild-to-moderate/severe intellectual impairment. Compared to developmental age, patients showed cognitive deficits in attention, (non)verbal memory and verbal fluency. Furthermore, dysarthria and weak oral-motor performance was found. Language was one of the patients' assets. In contrast to the severe deterioration of motor functioning in A-T, cognitive and language functioning appeared to level off with a typical profile of neuropsychological strengths and weaknesses. Based on our experiences with A-T, suggestions are made to determine a valid assessment of the cognitive and speech-language manifestations.

  19. Breast cancer risk in ataxia telangiectasia (AT) heterozygotes: haplotype study in French AT families

    Science.gov (United States)

    Janin, N; Andrieu, N; Ossian, K; Laugé, A; Croquette, M-F; Griscelli, C; Debré, M; Bressac-de-Paillerets, B; Aurias, A; Stoppa-Lyonnet, D

    1999-01-01

    Epidemiological studies in ataxia telangiectasia (AT) families have suggested that AT heterozygotes could have an increased cancer risk, especially breast cancer (BC) in women. It has also been suggested that an increased sensibility of AT heterozygotes to the effect of ionizing radiation could be responsible for the increased BC risk. BC relative risk (RR) estimation in AT heterozygotes within families ascertained through AT children is presented here. Family data collected included demographic characteristics, occurrence of cancers, past radiation exposures and blood samples. DNA samples were studied using seven ATM linked microsatellites markers allowing AT haplotypes reconstitution. The relative risk of BC was assessed using French estimated incidence rates. A significant increase risk of BC is found among obligate ATM heterozygotes with a point estimate of 3.32 (P = 0.002). BC relative risk calculated according to age is significantly increased among the obligate ATM heterozygotes female relatives with an age ≤ 44 years (RR = 4.55, P = 0.005). The BC relative risk is statistically borderline among the obligate ATM heterozygote female relatives with an age ≥ 45 years (RR = 2.48, P = 0.08). The estimated BC relative risk among ATM heterozygotes is consistent with previously published data. However, the increased risk is only a little higher than classical reproductive risk factors and similar to the risk associated with a first-degree relative affected by BC. © 1999 Cancer Research Campaign PMID:10362113

  20. Ocular findings in Norwegian patients with ataxia-telangiectasia: a 5 year prospective cohort study.

    Science.gov (United States)

    Riise, Ruth; Ygge, Jan; Lindman, Carl; Stray-Pedersen, Asbjørg; Bek, Toke; Rødningen, Olaug Kristin; Heiberg, Arvid

    2007-08-01

    To describe the outcome of ophthalmologic examination of 10 Norwegian children with ataxia-telangiectasia (AT) followed through 5 years. Ten Norwegian patients with AT aged 2-22 years (three females, seven males) were examined. The diagnosis was confirmed clinically as well as with molecular genetic studies. Conventional ophthalmologic examination was performed and supplemented by photographs of the conjunctiva, video recordings and registration of eye motility in five consecutive years. Additionally conjunctival biopsies were performed at the end of the follow-up period. General ataxia was usually detected when the child started to walk. All children over the age of 4 years had abnormal saccade movements, a form of ocular motor apraxia. Conjunctival telangiectasias were mostly visible at 4-5 years, primarily within the palpebral fissure. Immunohistochemical examination of conjunctival biopsies showed an increased number of cross-sections of blood vessels and neurons surrounded by glial tissue. There was a tendency to slightly earlier onset of conjunctival telangiectasias in the patients homozygous for a founder mutation compared with the other patients. The diagnosis of AT can be supported at preschool age by the onset of ocular motor apraxia and conjunctival telangiectasias. The findings become more prominent with age. The conjunctival telangiectasias seem to appear slightly earlier in the patients who are homozygous for a Norwegian founder mutation than in the rest of the patients.

  1. Safety and caregiver satisfaction with gastrostomy in patients with Ataxia Telangiectasia

    Directory of Open Access Journals (Sweden)

    Crawford Thomas O

    2011-05-01

    Full Text Available Abstract Background Ataxia Telangiectasia (A-T is a rare monogenetic neurodegenerative disease with pulmonary, nutritional, and dysphagic complications. Gastrostomy tube (GT feedings are commonly recommended to manage these co-morbidities. In general, outcomes of GT placement in patients with progressive diseases that develop during childhood are not well characterized. The primary purposes of this study were to determine whether GT placement in patients with A-T would be tolerated and associated with caregiver satisfaction. Methods We completed a retrospective review of 175 patients who visited the A-T Children's Center at Johns Hopkins Hospital from 2001 through 2008, and identified 28 patients with A-T (19 males, 9 females who underwent GT placement for non-palliative reasons. Information was obtained from medical records, interviews with primary health care providers, and 24 (83% caregivers of patients with GT's who responded to survey requests. Results Twenty-five (89% patients tolerated GT placement and were a median of 5.0 (0.4-12.6 years post GT placement at the time of this investigation. Three (11% patients died within one month of GT placement. In comparison to patients who tolerated GT placement, patients with early mortality were older when GT's were placed (median 24.9 vs. 12.3 years, p = 0.006 and had developed a combination of dysphagia, nutritional, and respiratory problems. Caregivers of patients tolerating GT placement reported significant improvements in mealtime satisfaction and participation in daily activities. Conclusions GT placement can be well tolerated and associated with easier mealtimes in patients with A-T when feeding tubes are placed at young ages. Patients with childhood onset of disorders with predictable progression of the disease process and impaired swallowing may benefit from early versus late placement of feeding tubes.

  2. Quantitative evaluation of brain involvement in ataxia telangiectasia by diffusion weighted MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Firat, Ahmet Kemal [Inonu University Medical Faculty, Turgut Ozal Medical Center, Department of Radiology, Malatya 44280 (Turkey); Karakas, Hakki Muammer [Inonu University Medical Faculty, Turgut Ozal Medical Center, Department of Radiology, Malatya 44280 (Turkey)]. E-mail: hkarakas@inonu.edu.tr; Firat, Yezdan [Inonu University Medical Faculty, Turgut Ozal Medical Center, Department of Otorhinolaryngology, Malatya (Turkey); Yakinci, Cengiz [Inonu University Medical Faculty, Turgut Ozal Medical Center, Department of Pediatrics, Malatya (Turkey)

    2005-11-01

    Objective: To evaluate the value of diffusion weighted imaging (DWI) in diagnosing ataxia telangiectasia (AT) and to investigate the spatial distribution of cerebral microstructural changes caused by the disease. Methods: Six AT patients (9-13 years) and nine healthy control subjects were examined on 1.5 T scanner. In addition to conventional MR images, DWI were performed with a fat suppressed, multishot spin echo EPI sequence using B values of 0, 500 and 1000 s/mm{sup 2}. Mean ADC values were measured from 16 different supra and infratentorial location. The difference between controls and AT patients regarding ADC values, and the accuracy, sensitivity and specificity of them in discrimination were analyzed with t-tests, logistic regression analysis, ANOVA and ROC curves. Results: Conventional images of the controls were normal. In AT patients, the only conventional MR abnormality was cerebellar atrophy. The difference between both groups regarding mean ADC values was not significant for any of the cerebral structures. In contrary to cerebrum, cerebellar mean ADC values of patients and controls were statistically different (p < 0.011-0.0001). Patients and controls were classified with 100% accuracy using ADC values of cerebellar white matter and cortex together (p < 0.016). The cut-off ADC value (0.699 mm{sup 2}/s) for middle cerebellar cortex had produced highest (100%) sensitivity and specificity. There was a difference between superior, middle and inferior cerebellar cortex regarding ADC values (p < 0.026). Superior cerebellar cortex (0.987 {+-} 0.1956 mm{sup 2}/s) had higher ADC values than the middle and inferior cerebellar cortex. Conclusion: DWI provides a supplementary and objective imaging finding in AT. This finding is highly accurate in the radiological discrimination of healthy subjects and AT. Our findings also implicate that AT causes a diffuse atrophy and mostly affects superior part of the cortex.

  3. Is age a risk factor for liver disease and metabolic alterations in ataxia Telangiectasia patients?

    Science.gov (United States)

    Paulino, Talita Lemos; Rafael, Marina Neto; Hix, Sonia; Shigueoka, David Carlos; Ajzen, Sergio Aron; Kochi, Cristiane; Suano-Souza, Fabíola Isabel; da Silva, Rosangela; Costa-Carvalho, Beatriz T; Sarni, Roseli O S

    2017-08-04

    Ataxia telangiectasia (A-T) is a neurodegenerative disease that leads to mitochondrial dysfunction and oxidative stress. Insulin resistance (IR), type 2 diabetes and the risk for development of cardiovascular disease was recently associated as an extended phenotype of the disease. We aimed to assess IR; liver involvement; carotid intima-media thickness (cIMT) and metabolic alterations associated to cardiovascular risk in A-T patients, and relate them with age. Glucose metabolism alterations were found in 54.6% of the patients. Hepatic steatosis was diagnosed in 11/17 (64.7%) A-T patients. AST/ALT ratio > 1 was observed in 10/17 (58.8%). A strong positive correlation was observed between insulin sum concentrations with ALT (r = 0.782, p < 0.004) and age (r = 0.818, p = 0.002). Dyslipidemia was observed in 55.5% of the patients. The apolipoprotein (Apo-B)/ApoA-I ratio (r = 0.619; p < 0.01), LDL/HDL-c (r = 0.490; p < 0.05) and the Apo-B levels (r = 0.545; p < 0.05) were positively correlated to cIMT. Metabolic disorders implicated in cardiovascular and liver diseases are frequently observed in adolescent A-T patients and those tend to get worse as they become older. Therefore, nutritional intervention and the use of drugs may be necessary.

  4. Ataxia Telangiectasia-Mutated (ATMPolymorphisms and Risk of Lung Cancer in a Chinese Population

    Directory of Open Access Journals (Sweden)

    Ajay A. Myneni

    2017-06-01

    Full Text Available BackgroundThe ataxia telangiectasia-mutated (ATM gene has a key role in DNA repair including activation and stabilization of p53, which implicates the importance of ATM polymorphisms in the development of cancer. This study aims to investigate the association of two ATM single-nucleotide polymorphisms (SNPs with lung cancer, as well as their potential interaction with p53 gene and other known risk factors of lung cancer.MethodsA population-based case–control study was conducted in Taiyuan city, China with 399 cases and 466 controls matched on the distribution of age and sex of cases. The two ATM gene SNPs, ATMrs227060 and ATMrs228589 as well as p53 gene SNP, p53rs1042522 were genotyped using Sequenom platform. Unconditional logistic regression models were used to estimate crude and adjusted odds ratios (aOR and 95% confidence intervals (CIs. Adjusted models controlled for age, sex, and smoking status.ResultsThe study showed that TT genotype of ATMrs227060 (aOR = 1.58, 95% CI: 1.06–2.35 and AA genotype of ATMrs228589 were significantly associated with lung cancer (aOR = 1.50, 95% CI: 1.08–2.08 in a recessive model. Additionally, carrying variant genotypes of ATMrs227060 (TT, ATMrs228589 (AA, and p53rs1042522 (CC concomitantly was associated with much higher risk (aOR = 3.68, 95% CI: 1.43–9.45 of lung cancer than carrying variant genotypes of any one of the above three SNPs. We also found multiplicative and additive interaction between tea drinking and ATMrs227060 in association with lung cancer.ConclusionThis study indicates that ATM gene variants might be associated with development of lung cancer in Chinese population. These results need to be validated in larger and different population samples.

  5. Telomere length, ATM mutation status and cancer risk in Ataxia-Telangiectasia families.

    Science.gov (United States)

    Renault, Anne-Laure; Mebirouk, Noura; Cavaciuti, Eve; Le Gal, Dorothée; Lecarpentier, Julie; d'Enghien, Catherine Dubois; Laugé, Anthony; Dondon, Marie-Gabrielle; Labbé, Martine; Lesca, Gaetan; Leroux, Dominique; Gladieff, Laurence; Adenis, Claude; Faivre, Laurence; Gilbert-Dussardier, Brigitte; Lortholary, Alain; Fricker, Jean-Pierre; Dahan, Karin; Bay, Jacques-Olivier; Longy, Michel; Buecher, Bruno; Janin, Nicolas; Zattara, Hélène; Berthet, Pascaline; Combès, Audrey; Coupier, Isabelle; Hall, Janet; Stoppa-Lyonnet, Dominique; Andrieu, Nadine; Lesueur, Fabienne

    2017-10-01

    Recent studies have linked constitutive telomere length (TL) to aging-related diseases including cancer at different sites. ATM participates in the signaling of telomere erosion, and inherited mutations in ATM have been associated with increased risk of cancer, particularly breast cancer. The goal of this study was to investigate whether carriage of an ATM mutation and TL interplay to modify cancer risk in ataxia-telangiectasia (A-T) families.The study population consisted of 284 heterozygous ATM mutation carriers (HetAT) and 174 non-carriers (non-HetAT) from 103 A-T families. Forty-eight HetAT and 14 non-HetAT individuals had cancer, among them 25 HetAT and 6 non-HetAT were diagnosed after blood sample collection. We measured mean TL using a quantitative PCR assay and genotyped seven single-nucleotide polymorphisms (SNPs) recurrently associated with TL in large population-based studies.HetAT individuals were at increased risk of cancer (OR = 2.3, 95%CI = 1.2-4.4, P = 0.01), and particularly of breast cancer for women (OR = 2.9, 95%CI = 1.2-7.1, P = 0.02), in comparison to their non-HetAT relatives. HetAT individuals had longer telomeres than non-HetAT individuals (P = 0.0008) but TL was not associated with cancer risk, and no significant interaction was observed between ATM mutation status and TL. Furthermore, rs9257445 (ZNF311) was associated with TL in HetAT subjects and rs6060627 (BCL2L1) modified cancer risk in HetAT and non-HetAT women.Our findings suggest that carriage of an ATM mutation impacts on the age-related TL shortening and that TL per se is not related to cancer risk in ATM carriers. TL measurement alone is not a good marker for predicting cancer risk in A-T families. © The Author 2017. Published by Oxford University Press.

  6. Ataxia-telangiectasia-mutated (ATM) and NBS1-dependent phosphorylation of Chk1 on Ser-317 in response to ionizing radiation

    DEFF Research Database (Denmark)

    Gatei, Magtouf; Sloper, Katie; Sørensen, Claus Storgaard

    2003-01-01

    In mammals, the ATM (ataxia-telangiectasia-mutated) and ATR (ATM and Rad3-related) protein kinases function as critical regulators of the cellular DNA damage response. The checkpoint functions of ATR and ATM are mediated, in part, by a pair of checkpoint effector kinases termed Chk1 and Chk2...

  7. Ataxia Telangiectasia-Mutated (ATM) kinase activity is regulated by ATP-driven conformational changes in the Mre11/Rad50/Nbs1 (MRN) complex

    NARCIS (Netherlands)

    J.-H. Lee (Ji-Hoon); M.R. Mand (Michael); R.A. Deshpande (Rajashree); E. Kinoshita (Eri); S.-H. Yang (Soo-Hyun); C. Wyman (Claire); T.T. Paull

    2013-01-01

    textabstractThe Ataxia Telangiectasia-Mutated (ATM) protein kinase is recruited to sites of double-strand DNA breaks by the Mre11/Rad50/Nbs1 (MRN) complex, which also facilitates ATM monomerization and activation. MRN exists in at least two distinct conformational states, dependent on ATP binding

  8. A YAC contig spanning the ataxia-telangiectasia locus (groups A and C) at 11q22-q23

    Energy Technology Data Exchange (ETDEWEB)

    Rotman, G.; Savitsky, K.; Ziv, Y. [Tel Aviv Univ. Ramat Aviv (Israel)] [and others

    1994-11-15

    Ataxia-telangiectasia (A-T) is an autosomal recessive disease involving cerebellar degeneration, immunodeficiency, cancer predisposition, chromosomal instability and radiosensitivity. A-T is heterogeneous, and the majority of A-T cases are associated with two complementation groups, A and C. The ATA and ATC loci are closely linked at chromosome 11q22-q23. Recombination mapping and linkage disequilibrium analysis have confined both loci between the markers D11S1817 and D11S927. Construction of this contig was expedited by prior generation of a region-specific ICRF sublibrary using Alu-PCR products derived from a radiation hybrid. The contig was expanded further by screening the libraries with Alu-PCR products derived from YAC clones and with STSs from YAC ends. YAC clones were aligned by fingerprinting with moderately repetitive probes. 56 refs., 5 figs., 1 tab.

  9. Imaging study of lymphoreticular tumor development in ataxia-telangiectasia and Nijmegen breakage syndrome; Estudio por imagen del desarrollo de tumores linforreticulares en la ataxia telangiectasia y el sindrome de Nijmegen

    Energy Technology Data Exchange (ETDEWEB)

    Martinez-Leon, M. I.; Ceres-Ruiz, L.; Cuesta, M. A.; Garcia-Martin, F. J. [Hospital Materno-Infantil C.H.U. Carlos Haya. Malaga (Spain)

    2003-07-01

    Ataxia-telangiectasia (AT), or Louis-Bar syndrome, is an autosomal recessive illness characterized by progressive cerebellar ataxia, oculo-cutaneous telangiectasia, immunodeficiency combined with susceptibility to sinopulmonary infections and high incidence of neoplastic development. Nijmegen breakage syndrome (NBS) is a variant of AT, is also an autosomal recessive illness that presents cerebellar ataxia, as well as combined immunodeficiency and a tendency toward tumor development. Contrary to Louis-Bar syndrome, it doesn't present telangiectasia and exhibits a characteristics phenotype (short stature, bird-like face and microcephaly). Both entities are classified as syndrome of chromosomal instability or chromosomal fragility, a group which also includes Bloom syndrome and Fanconi anemia. All of these show an increase in the frequency of neoplastic pathologies, mainly lymphoid tumors. We present three patients,two with AT and one with NBS, who developed different lymphoma types in the course of the illness. We highlight the most outstanding aspects from a clinical-radiological point of view. (Author) 17 refs.

  10. Ataxia telangiectasia: un desorden multisistémico con inestabilidad cromosómica y predisposición al cáncer

    OpenAIRE

    M. Guzmán

    1994-01-01

    El síndrome de ataxia telangiectasia (A-T) fue descrito por Syllaba y Henneren 1926 y redescrito en 1941 por Louis Bar (1,2). Es una entidad autosómica recesiva que afecta a hombres y mujeres en igual proporción. La expresividad del gen A-Tes variable y la incidencia familiares alta (1,2). Se ha estimado que la frecuencia de individuos homocigotos (afectados) para el gen A-T, es de 1/40.000 nacidos vivos y 1% de la población general serían heterocigotos (portadores). Sin embargo, la incidenci...

  11. Modest increased sensitivity to radiation oncogenesis in ATM heterozygous versus wild-type mammalian cells

    Science.gov (United States)

    Smilenov, L. B.; Brenner, D. J.; Hall, E. J.

    2001-01-01

    Subpopulations that are genetically predisposed to radiation-induced cancer could have significant public health consequences. Individuals homozygous for null mutations at the ataxia telangiectasia gene are indeed highly radiosensitive, but their numbers are very small. Ataxia Telangiectasia heterozygotes (1-2% of the population) have been associated with somewhat increased radiosensitivity for some end points, but none directly related to carcinogenesis. Here, intralitter comparisons between wild-type mouse embryo fibroblasts and mouse embryo fibroblasts carrying ataxia telangiectasia mutated (ATM) null mutation indicate that the heterozygous cells are more sensitive to radiation oncogenesis than their normal, litter-matched, counterparts. From these data we suggest that Ataxia Telangiectasia heterozygotes could indeed represent a societally-significant radiosensitive human subpopulation.

  12. Activation of eNOS in endothelial cells exposed to ionizing radiation involves components of the DNA damage response pathway

    Energy Technology Data Exchange (ETDEWEB)

    Nagane, Masaki; Yasui, Hironobu; Sakai, Yuri; Yamamori, Tohru [Laboratory of Radiation Biology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan); Niwa, Koichi [Laboratory of Biochemistry, Department of Food and Cosmetic Science, Faculty of Bioindustry, Tokyo University of Agriculture, Abashiri 099-2493 (Japan); Hattori, Yuichi [Department of Molecular and Medical Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Kondo, Takashi [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Inanami, Osamu, E-mail: inanami@vetmed.hokudai.ac.jp [Laboratory of Radiation Biology, Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818 (Japan)

    2015-01-02

    Highlights: • eNOS activity is increased in BAECs exposed to X-rays. • ATM is involved in this increased eNOS activity. • HSP90 modulates the radiation-induced activation of ATM and eNOS. - Abstract: In this study, the involvement of ataxia telangiectasia mutated (ATM) kinase and heat shock protein 90 (HSP90) in endothelial nitric oxide synthase (eNOS) activation was investigated in X-irradiated bovine aortic endothelial cells. The activity of nitric oxide synthase (NOS) and the phosphorylation of serine 1179 of eNOS (eNOS-Ser1179) were significantly increased in irradiated cells. The radiation-induced increases in NOS activity and eNOS-Ser1179 phosphorylation levels were significantly reduced by treatment with either an ATM inhibitor (Ku-60019) or an HSP90 inhibitor (geldanamycin). Geldanamycin was furthermore found to suppress the radiation-induced phosphorylation of ATM-Ser1181. Our results indicate that the radiation-induced eNOS activation in bovine aortic endothelial cells is regulated by ATM and HSP90.

  13. Ataxia Telangiectasia-Mutated (ATM) Kinase Activity Is Regulated by ATP-driven Conformational Changes in the Mre11/Rad50/Nbs1 (MRN) Complex*

    Science.gov (United States)

    Lee, Ji-Hoon; Mand, Michael R.; Deshpande, Rajashree A.; Kinoshita, Eri; Yang, Soo-Hyun; Wyman, Claire; Paull, Tanya T.

    2013-01-01

    The Ataxia Telangiectasia-Mutated (ATM) protein kinase is recruited to sites of double-strand DNA breaks by the Mre11/Rad50/Nbs1 (MRN) complex, which also facilitates ATM monomerization and activation. MRN exists in at least two distinct conformational states, dependent on ATP binding and hydrolysis by the Rad50 protein. Here we use an ATP analog-sensitive form of ATM to determine that ATP binding, but not hydrolysis, by Rad50 is essential for MRN stimulation of ATM. Mre11 nuclease activity is dispensable, although some mutations in the Mre11 catalytic domain block ATM activation independent of nuclease function, as does the mirin compound. The coiled-coil domains of Rad50 are important for the DNA binding ability of MRN and are essential for ATM activation, but loss of the zinc hook connection can be substituted by higher levels of the complex. Nbs1 binds to the “closed” form of the MR complex, promoted by the zinc hook and by ATP binding. Thus the primary role of the hook is to tether Rad50 monomers together, promoting the association of the Rad50 catalytic domains into a form that binds ATP and also binds Nbs1. Collectively, these results show that the ATP-bound form of MRN is the critical conformation for ATM activation. PMID:23525106

  14. Ataxia telangiectasia-mutated (ATM) kinase activity is regulated by ATP-driven conformational changes in the Mre11/Rad50/Nbs1 (MRN) complex.

    Science.gov (United States)

    Lee, Ji-Hoon; Mand, Michael R; Deshpande, Rajashree A; Kinoshita, Eri; Yang, Soo-Hyun; Wyman, Claire; Paull, Tanya T

    2013-05-03

    The Ataxia Telangiectasia-Mutated (ATM) protein kinase is recruited to sites of double-strand DNA breaks by the Mre11/Rad50/Nbs1 (MRN) complex, which also facilitates ATM monomerization and activation. MRN exists in at least two distinct conformational states, dependent on ATP binding and hydrolysis by the Rad50 protein. Here we use an ATP analog-sensitive form of ATM to determine that ATP binding, but not hydrolysis, by Rad50 is essential for MRN stimulation of ATM. Mre11 nuclease activity is dispensable, although some mutations in the Mre11 catalytic domain block ATM activation independent of nuclease function, as does the mirin compound. The coiled-coil domains of Rad50 are important for the DNA binding ability of MRN and are essential for ATM activation, but loss of the zinc hook connection can be substituted by higher levels of the complex. Nbs1 binds to the "closed" form of the MR complex, promoted by the zinc hook and by ATP binding. Thus the primary role of the hook is to tether Rad50 monomers together, promoting the association of the Rad50 catalytic domains into a form that binds ATP and also binds Nbs1. Collectively, these results show that the ATP-bound form of MRN is the critical conformation for ATM activation.

  15. Ataxia telangiectasia: un desorden multisistémico con inestabilidad cromosómica y predisposición al cáncer

    Directory of Open Access Journals (Sweden)

    M. Guzmán

    1994-12-01

    Full Text Available El síndrome de ataxia telangiectasia (A-T fue descrito por Syllaba y Henneren 1926 y redescrito en 1941 por Louis Bar (1,2. Es una entidad autosómica recesiva que afecta a hombres y mujeres en igual proporción. La expresividad del gen A-Tes variable y la incidencia familiares alta (1,2. Se ha estimado que la frecuencia de individuos homocigotos (afectados para el gen A-T, es de 1/40.000 nacidos vivos y 1% de la población general serían heterocigotos (portadores. Sin embargo, la incidencia en la población se puede incrementar ya que los homocigotos pueden tener descendencia. Es así como se estima que en la población blanca de Estados Unidos los heterocigotos podrían llegar a constituir el 1,4% de la población (2,3. Otros autores estiman una frecuencia de heterocigotos mucho más alta entre 0,68% y 7,7% de la población (4.

  16. Chromosomal Aberrations in Normal and AT Cells Exposed to High Dose of Low Dose Rate Irradiation

    Science.gov (United States)

    Kawata, T.; Shigematsu, N.; Kawaguchi, O.; Liu, C.; Furusawa, Y.; Hirayama, R.; George, K.; Cucinotta, F.

    2011-01-01

    Ataxia telangiectasia (A-T) is a human autosomally recessive syndrome characterized by cerebellar ataxia, telangiectases, immune dysfunction, and genomic instability, and high rate of cancer incidence. A-T cell lines are abnormally sensitive to agents that induce DNA double strand breaks, including ionizing radiation. The diverse clinical features in individuals affected by A-T and the complex cellular phenotypes are all linked to the functional inactivation of a single gene (AT mutated). It is well known that cells deficient in ATM show increased yields of both simple and complex chromosomal aberrations after high-dose-rate irradiation, but, less is known on how cells respond to low-dose-rate irradiation. It has been shown that AT cells contain a large number of unrejoined breaks after both low-dose-rate irradiation and high-dose-rate irradiation, however sensitivity for chromosomal aberrations at low-dose-rate are less often studied. To study how AT cells respond to low-dose-rate irradiation, we exposed confluent normal and AT fibroblast cells to up to 3 Gy of gamma-irradiation at a dose rate of 0.5 Gy/day and analyzed chromosomal aberrations in G0 using fusion PCC (Premature Chromosomal Condensation) technique. Giemsa staining showed that 1 Gy induces around 0.36 unrejoined fragments per cell in normal cells and around 1.35 fragments in AT cells, whereas 3Gy induces around 0.65 fragments in normal cells and around 3.3 fragments in AT cells. This result indicates that AT cells can rejoin breaks less effectively in G0 phase of the cell cycle? compared to normal cells. We also analyzed chromosomal exchanges in normal and AT cells after exposure to 3 Gy of low-dose-rate rays using a combination of G0 PCC and FISH techniques. Misrejoining was detected in the AT cells only? When cells irradiated with 3 Gy were subcultured and G2 chromosomal aberrations were analyzed using calyculin-A induced PCC technique, the yield of unrejoined breaks decreased in both normal and AT

  17. The defect in the AT-like hamster cell mutants is complemented by mouse chromosome 9 but not by any of the human chromosomes

    NARCIS (Netherlands)

    W. Jongmans; G. Verhaegh (Gerald); N.G.J. Jaspers (Nicolaas); P. Demant (Peter); A.T. Natarajan; Y. Shiloh (Yosef); M. Oshimura (Mitsuo); E.J. Stanbridge (Eric); R.S. Athwal (Raghbir); A.P. Cuthbert (Andrew); R.F. Newbold (Robert); P.H.M. Lohmann (Paul); M.Z. Zdzienicka (Malgorzata)

    1996-01-01

    textabstractX-ray-sensitive Chinese hamster V79 cells mutants, V-C4, V-E5 and V-G8, show an abnormal response to X-ray-induced DNA damage. Like ataxia telangiectasia (AT) cells, they display increased cell killing, chromosomal instability and a diminished inhibition of DNA synthesis following

  18. Review of Chromium (VI) Apoptosis, Cell-Cycle-Arrest, and Carcinogenesis

    Science.gov (United States)

    Chiu, A; Shi, J; Lee, WKP; Hill, R; Wakeman, TP; Katz, A; Xu, B; Dalal, NS; Robertson, JD; Chen, C; Chiu, N; Donehower, L

    2014-01-01

    Hexavalent chromium combines with glutathione in chloride intracellular channel carrier to form tetravalent and pentavelent chromium in plasma and organelle membranes. It also combines with NADH/NADPH to form pentavalent chromium in mitochondria. Tetravalent- and pentavalent- chromium (directly and indirectly) mediated DNA double strand breaks activate DNA damage signaling sensors: DNA-dependent-protein-kinase signals p53-dependent intrinsic mitochorndrial apoptosis, and ataxia-telangiectasia-mutated and ataxia-telangiectasia-Rad3-related signal cell-arrest for DNA repair. Tetravalent chromium may be the most potent species since it causes DNA breaks and somatic recombination, but not apoptosis. Upon further failure of apoptosis and senescence/DNA-repair, damaged cells may become immortal with loss-of-heterozygosity and genetic plasticity. PMID:20859824

  19. Methacryloxylethyl Cetyl Ammonium Chloride Induces DNA Damage and Apoptosis in Human Dental Pulp Cells via Generation of Oxidative Stress.

    Science.gov (United States)

    Jiao, Yang; Ma, Sai; Wang, Yirong; Li, Jing; Shan, Lequn; Sun, Jinlong; Chen, Jihua

    2016-01-01

    The polymerizable antibacterial monomer methacryloxylethyl cetyl ammonium chloride (DMAE-CB) has provided an effective strategy to combat dental caries. However, the application of such material raises the question about the biological safety and the question remains open. The mechanism of this toxic action, however, is not yet clearly understood. The present study aims at providing novel insight into the possible causal link between cellular oxidative stress and DNA damage, as well as apoptosis in human dental pulp cells exposed to DMAE-CB. The enhanced formation of reactive oxygen species and depletion of glutathione, as well as differential changes in activities of superoxide dismutase, glutathione peroxidase, and catalase in DMAE-CB-treated cells indicated oxidative stress. By using substances that can alter GSH synthesis, we found that GSH was the key component in the regulation of cell response towards oxidative stress induced by DMAE-CB. The increase in oxidative stress-sensitive 8-Oxo-2'-deoxyguanosine (8-OHdG) content, formation of γ-H2AX and cell cycle G1 phase arrest indicated that DNA damage occurred as a result of the interaction between DNA base and ROS beyond the capacities of antioxidant mechanisms in cells exposed to DMAE-CB. Such oxidative DNA damage thus triggers the activation of ataxia telangiectasia-mutated (ATM) signaling, the intrinsic apoptotic pathway, and destruction of mitochondrial morphology and function.

  20. Inhibition of DNA-PKcs enhances radiosensitivity and increases the levels of ATM and ATR in NSCLC cells exposed to carbon ion irradiation.

    Science.gov (United States)

    Yang, Lina; Liu, Yuanyuan; Sun, Chao; Yang, Xinrui; Yang, Zhen; Ran, Juntao; Zhang, Qiuning; Zhang, Hong; Wang, Xinyu; Wang, Xiaohu

    2015-11-01

    Non-small cell lung cancer (NSCLC) exhibits radioresistance to conventional rays, due to its DNA damage repair systems. NSCLC may potentially be sensitized to radiation treatment by reducing those factors that continuously enhance the repair of damaged DNA. In the present study, normal lung fibroblast MRC-5 and lung cancer A549 cells were treated with NU7026 and CGK733, which are inhibitors of the DNA-dependent protein kinase catalytic subunit (PKcs) and ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related (ATR), respectively, followed by exposure to X-rays and carbon ion irradiation. The cytotoxic activity, cell survival rate, DNA damage repair ability, cell cycle arrest and apoptosis rate of the treated cells were analyzed with MTT assay, colony formation assay, immunofluorescence and flow cytometry, respectively. The transcription and translation levels of the ATM, ATR and DNA-PKcs genes were detected by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. The results indicated that the radiosensitivity and DNA repair ability of A549 cells were reduced, and the percentages of apoptotic cells and those arrested at the G2/M phase of the cell cycle were significantly increased, following ionizing radiation with inhibitor-pretreatment. The expression levels of ATM, ATR, DNA-PKcs and phosphorylated histone H2AX, a biomarker for DNA double-strand breaks, were all upregulated at the transcriptional or translational level in A549 cells treated with carbon ion irradiation, compared with the control and X-rays-treated cells. In addition, the treatment with 5-50 µM NU7026 or CGK733 did not produce any obvious cytotoxicity in MRC-5 cells, and the effect of the DNA-PKcs-inhibitor on enhancing the radiosensitivity of A549 cells was stronger than that observed for the ATM and ATR-inhibitor. These findings demonstrated a minor role for ATM and ATR in radiation-induced cell death, since the upregulation of

  1. Rapamycin enhances lytic replication of Epstein-Barr virus in gastric carcinoma cells by increasing the transcriptional activities of immediate-early lytic promoters.

    Science.gov (United States)

    Wang, Man; Wu, Wei; Zhang, Yinfeng; Yao, Guoliang; Gu, Bianli

    2017-11-21

    Epstein-Barr virus (EBV), a human herpesvirus, is linked to both epithelial and lymphoid malignancies. Induction of EBV reactivation is a potential therapeutic strategy for EBV-associated tumors. In this study, we assessed the effects of rapamycin on EBV reactivation in gastric carcinoma cells. We found that rapamycin upregulated expression of EBV lytic proteins and increased the viral proliferation triggered by the EBV lytic inducer sodium butyrate. Reverse transcription-qPCR, luciferase activity assays, chromatin immunoprecipitation and western blotting were employed to explore the mechanism by which rapamycin promotes EBV reactivation. Our results showed that rapamycin treatment resulted in increased mRNA levels of EBV immediate-early genes. Rapamycin also enhanced the transcriptional activities of the EBV immediate-early lytic promoters Zp and Rp by strengthening Sp1 binding. Repression of the cellular ataxia telangiectasia-mutated/p53 pathway by siRNA-mediated knockdown of the ataxia telangiectasia-mutated gene significantly abrogated virus reactivation by rapamycin/sodium butyrate treatment, indicating that the ataxia telangiectasia-mutated/p53 pathway is involved in rapamycin-promoted EBV reactivation. Taken together, these findings demonstrate that rapamycin might have the potential to enhance the effectiveness of oncolytic viral therapies developed for EBV-associated malignancies. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Breast cancer stem cell-like cells are more sensitive to ionizing radiation than non-stem cells: role of ATM.

    Directory of Open Access Journals (Sweden)

    Seog-Young Kim

    Full Text Available There are contradictory observations about the different radiosensitivities of cancer stem cells and cancer non-stem cells. To resolve these contradictory observations, we studied radiosensitivities by employing breast cancer stem cell (CSC-like MDA-MB231 and MDA-MB453 cells as well as their corresponding non-stem cells. CSC-like cells proliferate without differentiating and have characteristics of tumor-initiating cells [1]. These cells were exposed to γ-rays (1.25-8.75 Gy and survival curves were determined by colony formation. A final slope, D(0, of the survival curve for each cell line was determined to measure radiosensitivity. The D(0 of CSC-like and non-stem MDA-MB-453 cells were 1.16 Gy and 1.55 Gy, respectively. Similar results were observed in MDA-MB-231 cells (0.94 Gy vs. 1.56 Gy. After determination of radiosensitivity, we investigated intrinsic cellular determinants which influence radiosensitivity including cell cycle distribution, free-radical scavengers and DNA repair. We observed that even though cell cycle status and antioxidant content may contribute to differential radiosensitivity, differential DNA repair capacity may be a greater determinant of radiosensitivity. Unlike non-stem cells, CSC-like cells have little/no sublethal damage repair, a low intracellular level of ataxia telangiectasia mutated (ATM and delay of γ-H2AX foci removal (DNA strand break repair. These results suggest that low DNA repair capacity is responsible for the high radiosensitivity of these CSC-like cells.

  3. MicroRNA-22 promotes cell survival upon UV radiation by repressing PTEN

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Guangyun [Department of Pathology and Laboratory Medicine, University of Tennessee Health Science Center, Memphis, TN (United States); Center for Adult Cancer Research, University of Tennessee Health Science Center, Memphis, TN (United States); Jilin Province Key Laboratory of Animal Embryo Engineering, Jilin University, Changchun (China); Shi, Yuling [Department of Pathology and Laboratory Medicine, University of Tennessee Health Science Center, Memphis, TN (United States); Center for Adult Cancer Research, University of Tennessee Health Science Center, Memphis, TN (United States); Wu, Zhao-Hui, E-mail: zwu6@uthsc.edu [Department of Pathology and Laboratory Medicine, University of Tennessee Health Science Center, Memphis, TN (United States); Center for Adult Cancer Research, University of Tennessee Health Science Center, Memphis, TN (United States)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer miR-22 is induced in cells treated with UV radiation. Black-Right-Pointing-Pointer ATM is required for miR-22 induction in response to UV. Black-Right-Pointing-Pointer miR-22 targets 3 Prime -UTR of PTEN to repress its expression in UV-treated cells. Black-Right-Pointing-Pointer Upregulated miR-22 inhibits apoptosis in cells exposed to UV. -- Abstract: DNA damage response upon UV radiation involves a complex network of cellular events required for maintaining the homeostasis and restoring genomic stability of the cells. As a new class of players involved in DNA damage response, the regulation and function of microRNAs in response to UV remain poorly understood. Here we show that UV radiation induces a significant increase of miR-22 expression, which appears to be dependent on the activation of DNA damage responding kinase ATM (ataxia telangiectasia mutated). Increased miR-22 expression may result from enhanced miR-22 maturation in cells exposed to UV. We further found that tumor suppressor gene phosphatase and tensin homolog (PTEN) expression was inversely correlated with miR-22 induction and UV-induced PTEN repression was attenuated by overexpression of a miR-22 inhibitor. Moreover, increased miR-22 expression significantly inhibited the activation of caspase signaling cascade, leading to enhanced cell survival upon UV radiation. Collectively, these results indicate that miR-22 is an important player in the cellular stress response upon UV radiation, which may promote cell survival via the repression of PTEN expression.

  4. Multispectral imaging flow cytometry reveals distinct frequencies of γ-H2AX foci induction in DNA double strand break repair defective human cell lines.

    Science.gov (United States)

    Bourton, Emma C; Plowman, Piers N; Zahir, Sheba Adam; Senguloglu, Gonul Ulus; Serrai, Hiba; Bottley, Graham; Parris, Christopher N

    2012-02-01

    The measurement of γ-H2AX foci induction in cells provides a sensitive and reliable method for the quantitation of DNA damage responses in a variety of cell types. Accurate and rapid methods to conduct such observations are desirable. In this study, we have employed the novel technique of multispectral imaging flow cytometry to compare the induction and repair of γ-H2AX foci in three human cell types with different capacities for the repair of DNA double strand breaks (DSB). A repair normal fibroblast cell line MRC5-SV1, a DSB repair defective ataxia telangiectasia (AT5BIVA) cell line, and a DNA-PKcs deficient cell line XP14BRneo17 were exposed to 2 Gy gamma radiation from a (60)Cobalt source. Thirty minutes following exposure, we observed a dramatic induction of foci in the nuclei of these cells. After 24 hrs, there was a predictable reduction on the number of foci in the MRC5-SV1 cells, consistent with the repair of DNA DSB. In the AT5BIVA cells, persistence of the foci over a 24-hr period was due to the failure in the repair of DNA DSB. However, in the DNA-PKcs defective cells (XP14BRneo17), we observed an intermediate retention of foci in the nuclei indicative of partial repair of DNA DSB. In summary, the application of imaging flow cytometry has permitted an evaluation of foci in a large number of cells (20,000) for each cell line at each time point. This provides a novel method to determine differences in repair kinetics between different cell types. We propose that imaging flow cytometry provides an alternative platform for accurate automated high through-put analysis of foci induction in a variety of cell types. Copyright © 2011 International Society for Advancement of Cytometry.

  5. Survey of radiosensitivity in a variety of human cell strains

    Energy Technology Data Exchange (ETDEWEB)

    Arlett, C.F.; Harcourt, S.A.

    1980-03-01

    Gamma-ray sensitivity for cell killing was assayed in 54 human cell strains, including some derived from individuals suffering from certain hereditary diseases. The overall range of Do values in this study was 38 to 180 rads, indicating a considerable range of variability in humans. The normal sensitivity was described by a range of Do values of 97 to 180 rads. All ten ataxia telangiectasia cell strains tested proved radiosensitive and gave a mean Do value of 57 +- 15 (S.E.) rads, and these represent the most radiosensitive human skin fibroblasts currently available. Representative cell strains from familial retinoblastoma, Fanconi's anemia, and Hutchinson-Gilford progeria occupied positions of intermediate sensitivity, as did one of two ataxia telangiectasia heterozygotes. Six xeroderma pigmentosum cell strains together with two Cockayne's syndrome cell strains (all known to be sensitive to ultraviolet light) fell into the normal range, indicating an absence of cross-sensitivity between ultraviolet light and gamma-irradiation.

  6. ATM-activated autotaxin (ATX) propagates inflammation and DNA damage in lung epithelial cells: a new mode of action for silica-induced DNA damage?

    Science.gov (United States)

    Zheng, Huiyuan; Högberg, Johan; Stenius, Ulla

    2017-12-07

    Silica exposure is a common risk factor for lung cancer. It has been claimed that key elements in cancer development are activation of inflammatory cells that indirectly induce DNA damage and proliferative stimuli in respiratory epithelial cells. We studied DNA damage induced by silica particles in respiratory epithelial cells and focused the role of the signaling enzyme autotaxin (ATX). A549 and 16 bronchial epithelial cells (16HBE) lung epithelial cells were exposed to silica particles. Reactive oxygen species (ROS), NOD-like receptor family pyrin domain containing-3 (NLRP3) inflammasome activation, ATX, ataxia telangiectasia mutated (ATM), and DNA damage (γH2AX, pCHK1, pCHK2, comet assay) were end points. Low doses of silica induced NLRP3 activation, DNA damage accumulation, and ATM phosphorylation. A novel finding was that ATM induced ATX generation and secretion. Not only silica but also rotenone, camptothecin and H2O2 activated ATX via ATM, suggesting that ATX is part of a generalized ATM response to double-strand breaks (DSBs). Surprisingly, ATX inhibition mitigated DNA damage accumulation at later time points (6-16 h), and ATX transfection caused NLRP3 activation and DNA damage. Furthermore, the product of ATX enzymatic activity, lysophosphatidic acid, recapitulated the effects of ATX transfection. These data indicate an ATM-ATX-dependent loop that propagates inflammation and DSB accumulation, making low doses of silica effective inducers of DSBs in epithelial cells. We conclude that an ATM-ATX axis interconnects DSBs with silica-induced inflammation and propagates these effects in epithelial cells. Further studies of this adverse outcome pathway may give an accurate assessment of the lowest doses of silica that causes cancer. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  7. ATM-Deficient Colorectal Cancer Cells Are Sensitive to the PARP Inhibitor Olaparib.

    Science.gov (United States)

    Wang, Chen; Jette, Nicholas; Moussienko, Daniel; Bebb, D Gwyn; Lees-Miller, Susan P

    2017-04-01

    The ataxia telangiectasia mutated (ATM) protein kinase plays a central role in the cellular response to DNA damage. Loss or inactivation of both copies of the ATM gene (ATM) leads to ataxia telangiectasia, a devastating childhood condition characterized by neurodegeneration, immune deficiencies, and cancer predisposition. ATM is also absent in approximately 40% of mantle cell lymphomas (MCLs), and we previously showed that MCL cell lines with loss of ATM are sensitive to poly-ADP ribose polymerase (PARP) inhibitors. Next-generation sequencing of patient tumors has revealed that ATM is altered in many human cancers including colorectal, lung, prostate, and breast. Here, we show that the colorectal cancer cell line SK-CO-1 lacks detectable ATM protein expression and is sensitive to the PARP inhibitor olaparib. Similarly, HCT116 colorectal cancer cells with shRNA depletion of ATM are sensitive to olaparib, and depletion of p53 enhances this sensitivity. Moreover, HCT116 cells are sensitive to olaparib in combination with the ATM inhibitor KU55933, and sensitivity is enhanced by deletion of p53. Together our studies suggest that PARP inhibitors may have potential for treating colorectal cancer with ATM dysfunction and/or colorectal cancer with mutation of p53 when combined with an ATM kinase inhibitor. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Punicalagin from pomegranate promotes human papillary thyroid carcinoma BCPAP cell death by triggering ATM-mediated DNA damage response.

    Science.gov (United States)

    Yao, Xin; Cheng, Xian; Zhang, Li; Yu, Huixin; Bao, Jiandong; Guan, Haixia; Lu, Rongrong

    2017-11-01

    Punicalagin (PUN), a component derived from pomegranate, is well known for its anticancer activity. Our previous work revealed that PUN induces autophagic cell death in papillary thyroid carcinoma cells. We hypothesized that PUN triggers DNA damage associated with cell death because DNA damage was reported as an inducer of autophagy. Our results showed that PUN treatment caused DNA breaks as evidenced by the significant enhancement in the phosphorylation of H2A.X. However, reactive oxygen species and DNA conformational alteration, 2 common inducing factors in DNA damage, were not involved in PUN-induced DNA damage. The phosphorylation of ataxia-telangiectasia mutated gene-encoded protein (ATM) but not ataxia telangiectasia and Rad3-related protein (ATR) was up-regulated in a time- and dosage-dependent manner after PUN treatment. KU-55933, an inhibitor of ATM, inhibited the phosphorylation of ATM induced by PUN and reversed the decreased cell viability caused by PUN. Thus, we demonstrated that PUN induces cell death of papillary thyroid carcinoma cells by triggering ATM-mediated DNA damage response, which provided novel mechanisms and potential targets for the better understanding of the anticancer actions of PUN. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Proteomic profiling of ATM kinase proficient and deficient cell lines upon blockage of proteasome activity.

    Science.gov (United States)

    Marzano, Valeria; Santini, Simonetta; Rossi, Claudia; Zucchelli, Mirco; D'Alessandro, Annamaria; Marchetti, Carlo; Mingardi, Michele; Stagni, Venturina; Barilà, Daniela; Urbani, Andrea

    2012-08-03

    Ataxia Telangiectasia Mutated (ATM) protein kinase is a key effector in the modulation of the functionality of some important stress responses, including DNA damage and oxidative stress response, and its deficiency is the hallmark of Ataxia Telangiectasia (A-T), a rare genetic disorder. ATM modulates the activity of hundreds of target proteins, essential for the correct balance between proliferation and cell death. The aim of this study is to evaluate the phenotypic adaptation at the protein level both in basal condition and in presence of proteasome blockage in order to identify the molecules whose level and stability are modulated through ATM expression. We pursued a comparative analysis of ATM deficient and proficient lymphoblastoid cells by label-free shotgun proteomic experiments comparing the panel of proteins differentially expressed. Through a non-supervised comparative bioinformatic analysis these data provided an insight on the functional role of ATM deficiency in cellular carbohydrate metabolism's regulation. This hypothesis has been demonstrated by targeted metabolic fingerprint analysis SRM (Selected Reaction Monitoring) on specific thermodynamic checkpoints of glycolysis. This article is part of a Special Issue entitled: Translational Proteomics. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Proteomic profiling of ATM kinase proficient and deficient cell lines upon blockage of proteasome activity☆

    Science.gov (United States)

    Marzano, Valeria; Santini, Simonetta; Rossi, Claudia; Zucchelli, Mirco; D'Alessandro, Annamaria; Marchetti, Carlo; Mingardi, Michele; Stagni, Venturina; Barilà, Daniela; Urbani, Andrea

    2012-01-01

    Ataxia Telangiectasia Mutated (ATM) protein kinase is a key effector in the modulation of the functionality of some important stress responses, including DNA damage and oxidative stress response, and its deficiency is the hallmark of Ataxia Telangiectasia (A-T), a rare genetic disorder. ATM modulates the activity of hundreds of target proteins, essential for the correct balance between proliferation and cell death. The aim of this study is to evaluate the phenotypic adaptation at the protein level both in basal condition and in presence of proteasome blockage in order to identify the molecules whose level and stability are modulated through ATM expression. We pursued a comparative analysis of ATM deficient and proficient lymphoblastoid cells by label-free shotgun proteomic experiments comparing the panel of proteins differentially expressed. Through a non-supervised comparative bioinformatic analysis these data provided an insight on the functional role of ATM deficiency in cellular carbohydrate metabolism's regulation. This hypothesis has been demonstrated by targeted metabolic fingerprint analysis SRM (Selected Reaction Monitoring) on specific thermodynamic checkpoints of glycolysis. This article is part of a Special Issue entitled: Translational Proteomics. PMID:22641158

  11. Radioresistance of chordoma cells is associated with the ATM/ATR pathway, in which RAD51 serves as an important downstream effector.

    Science.gov (United States)

    Zhang, Chao; Wang, Bing; Li, Lei; Li, Yawei; Li, Pengzhi; Lv, Guohua

    2017-09-01

    Surgery followed by radiotherapy is the standard treatment for chordomas, which are a rare but low-grade type of bone cancer arising from remnants of the embryonic notochord. However, disease recurrence following radiotherapy is common, most likely due to endogenous DNA repair mechanisms that promote cell survival upon radiation strikes. The ataxia telangiectasia mutated/ataxia telangiectasia mutated and Rad3 related (ATM/ATR)-mediated pathway has a critical role in DNA repair mechanisms; however, it has rarely been investigated in chordomas. In the present study, the expression of signal molecules related to the ATM/ATR pathway in chordoma tissues and adjacent normal tissues were initially examined using immunohistochemistry and western blot analysis. Chordoma U-CH1 and U-CH2 cells were subsequently used to investigate cell responses to ionizing radiation and the potential protective actions mediated by the ATM/ATR pathway. Phosphorylated (p)-ATM, p-ATR, γ-H2A histone family, member X (H2AX) and RAD51 were significantly upregulated in chordoma tissues relative to adjacent normal tissues (PATM, γ-H2AX and RAD51 expression in U-CH1 cells (PATM, p-ATR and RAD51 levels in U-CH2 cells (PATM/ATR pathway, in which RAD51 serves as an important downstream effector. Thus, RAD51 presents a promising therapeutic target for improving the outcome of radiotherapy treatment in chordomas.

  12. The BOSS Experiment of the EXPOSE-R2 Mission: Biofilms versus planktonic cells

    OpenAIRE

    Panitz, C.; Frösler, J.; Wingender, J.; Flemming, H.-C.; Rösch, P; Rettberg, P.

    2017-01-01

    In the BOSS experiment (biofilm organisms surfing space), which was performed in the context of the successfully finalized EXPOSE-R2 mission, an international consortium of scientists investigated the ability of a variety of organisms to survive in space and on Mars as a function of their life style. The question in focus is whether there are different strategies for individually living microorganisms (planktonic state) compared to a microbial consortium of the same cells (biofilm state) to c...

  13. Susceptibility of ATM-deficient pancreatic cancer cells to radiation.

    Science.gov (United States)

    Ayars, Michael; Eshleman, James; Goggins, Michael

    2017-05-19

    Ataxia telangiectasia mutated (ATM) is inactivated in a significant minority of pancreatic ductal adenocarcinomas and may be predictor of treatment response. We determined if ATM deficiency renders pancreatic cancer cells more sensitive to fractionated radiation or commonly used chemotherapeutics. ATM expression was knocked down in three pancreatic cancer cell lines using ATM-targeting shRNA. Isogenic cell lines were tested for sensitivity to several chemotherapeutic agents and radiation. DNA repair kinetics were analyzed in irradiated cells using the comet assay. We find that while rendering pancreatic cancer cells ATM-deficient did not significantly change their sensitivity to several chemotherapeutics, it did render them exquisitely sensitized to radiation. Pancreatic cancer ATM status may help predict response to radiotherapy.

  14. ATM regulation of IL-8 links oxidative stress to cancer cell migration and invasion.

    Science.gov (United States)

    Chen, Wei-Ta; Ebelt, Nancy D; Stracker, Travis H; Xhemalce, Blerta; Van Den Berg, Carla L; Miller, Kyle M

    2015-06-01

    Ataxia-telangiectasia mutated (ATM) protein kinase regulates the DNA damage response (DDR) and is associated with cancer suppression. Here we report a cancer-promoting role for ATM. ATM depletion in metastatic cancer cells reduced cell migration and invasion. Transcription analyses identified a gene network, including the chemokine IL-8, regulated by ATM. IL-8 expression required ATM and was regulated by oxidative stress. IL-8 was validated as an ATM target by its ability to rescue cell migration and invasion defects in ATM-depleted cells. Finally, ATM-depletion in human breast cancer cells reduced lung tumors in a mouse xenograft model and clinical data validated IL-8 in lung metastasis. These findings provide insights into how ATM activation by oxidative stress regulates IL-8 to sustain cell migration and invasion in cancer cells to promote metastatic potential. Thus, in addition to well-established roles in tumor suppression, these findings identify a role for ATM in tumor progression.

  15. miR-30a can inhibit DNA replication by targeting RPA1 thus slowing cancer cell proliferation.

    Science.gov (United States)

    Zou, Zhenyou; Ni, Mengjie; Zhang, Jing; Chen, Yongfeng; Ma, Hongyu; Qian, Shihan; Tang, Longhua; Tang, Jiamei; Yao, Hailun; Zhao, Chengbin; Lu, Xiongwen; Sun, Hongyang; Qian, Jue; Mao, Xiaoting; Lu, Xulin; Liu, Qun; Zen, Juping; Wu, Hanbing; Bao, Zhaosheng; Lin, Shudan; Sheng, Hongyu; Li, Yunlong; Liang, Yong; Chen, Zhiqiang; Zong, Dan

    2016-07-15

    Cell proliferation was inhibited following forced over-expression of miR-30a in the ovary cancer cell line A2780DX5 and the gastric cancer cell line SGC7901R. Interestingly, miR-30a targets the DNA replication protein RPA1, hinders the replication of DNA and induces DNA fragmentation. Furthermore, ataxia telangiectasia mutated (ATM) and checkpoint kinase 2 (CHK2) were phosphorylated after DNA damage, which induced p53 expression, thus triggering the S-phase checkpoint, arresting cell cycle progression and ultimately initiating cancer cell apoptosis. Therefore, forced miR-30a over-expression in cancer cells can be a potential way to inhibit tumour development. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

  16. Increased oxidative stress in AOA3 cells disturbs ATM-dependent DNA damage responses.

    Science.gov (United States)

    Kobayashi, Junya; Saito, Yuichiro; Okui, Michiyo; Miwa, Noriko; Komatsu, Kenshi

    2015-04-01

    Ataxia telangiectasia (AT) is caused by a mutation in the ataxia-telangiectasia-mutated (ATM) gene; the condition is associated with hyper-radiosensitivity, abnormal cell-cycle checkpoints, and genomic instability. AT patients also show cerebellar ataxia, possibly due to reactive oxygen species (ROS) sensitivity in neural cells. The ATM protein is a key regulator of the DNA damage response. Recently, several AT-like disorders have been reported. The genes responsible for them are predicted to encode proteins that interact with ATM in the DNA-damage response. Ataxia with oculomotor apraxia types 1-3 (AOA1, 2, and 3) result in a neurodegenerative and cellular phenotype similar to AT; however, the basis of this phenotypic similarity is unclear. Here, we show that the cells of AOA3 patients display aberrant ATM-dependent phosphorylation and apoptosis following γ-irradiation. The ATM-dependent response to H2O2 treatment was abrogated in AOA3 cells. Furthermore, AOA3 cells had reduced ATM activity. Our results suggest that the attenuated ATM-related response is caused by an increase in endogenous ROS in AOA3 cells. Pretreatment of cells with pyocyanin, which induces endogenous ROS production, abolished the ATM-dependent response. Moreover, AOA3 cells had decreased homologous recombination (HR) activity, and pyocyanin pretreatment reduced HR activity in HeLa cells. These results indicate that excess endogenous ROS represses the ATM-dependent cellular response and HR repair in AOA3 cells. Since the ATM-dependent cell-cycle checkpoint is an important block to carcinogenesis, such inactivation of ATM may lead to tumorigenesis as well as neurodegeneration. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. ATM mutants

    Indian Academy of Sciences (India)

    First page Back Continue Last page Graphics. ATM mutants. ATM (Ataxia Telangiectasia Mutated). AT2BE and AT5B1 cells – fibroblast cell lines from Ataxia telangiectasia patients. Deletion mutants expressing truncated ATM protein which is inactive. Have been used in studies looking at the role of ATM in DNA damage ...

  18. Study of ATM Phosphorylation by Cdk5 in Neuronal Cells.

    Science.gov (United States)

    She, Hua; Mao, Zixu

    2017-01-01

    The phosphatidylinositol-3-kinase-like kinase ATM (ataxia-telangiectasia mutated) plays a central role in coordinating the DNA damage responses including cell cycle checkpoint control, DNA repair, and apoptosis. Mutations of ATM cause a spectrum of defects ranging from neurodegeneration to cancer predisposition. We previously showed that Cdk5 (cyclin-dependent kinase 5) is activated by DNA damage and directly phosphorylates ATM at serine 794 in postmitotic neurons. Phosphorylation at serine 794 precedes and is required for ATM autophosphorylation at serine 1981, and activates ATM kinase activity. Cdk5-ATM pathway plays a crucial role in DNA damage-induced neuronal injury. This chapter describes protocols used in analyzing ATM phosphorylation by Cdk5 in CGNs (cerebellar granule neurons) and its effects on neuronal survival.

  19. Inflammatory Alteration of Human T Cells Exposed Continuously to Asbestos.

    Science.gov (United States)

    Kumagai-Takei, Naoko; Yamamoto, Shoko; Lee, Suni; Maeda, Megumi; Masuzzaki, Hidenori; Sada, Nagisa; Yu, Min; Yoshitome, Kei; Nishimura, Yasumitsu; Otsuki, Takemi

    2018-02-08

    Asbestos is a known carcinogen and exposure can lead to lung cancer and malignant mesothelioma. To examine the effects of asbestos fibers on human immune cells, the human T cell leukemia/lymphoma virus (HTLV)-1 immortalized human T cell line MT-2 was employed. Following continuous exposure to asbestos fibers for more than eight months, MT-2 sublines showed acquisition of resistance to asbestos-induced apoptosis with decreased death signals and increased surviving signals. These sublines showed various characteristics that suggested a reduction in anti-tumor immunity. On the other hand, inflammatory changes such as expression of MMP7, CXCR5, CXCL13 and CD44 was found to be markedly higher in sublines continuously exposed to asbestos compared with original MT-2 cells. All of these molecules contribute to lung inflammation, T and B cell interactions and connections between mesothelial cells and T cells. Thus, further investigation focusing on these molecules may shed light on the role of chronic inflammation caused by asbestos exposure and the occurrence of malignant mesothelioma. Finally, regarding peripheral T cells from healthy donors (HD) and asbestos-exposed patients with pleural plaque (PP) or malignant pleural mesothelioma (MPM), following stimulation of CD4+ T cells, T cells from MPM patients showed reduced potential of interferon (IFN)-γ expression. Moreover, levels of interleukin (IL)-6, one of the most important cytokines in chronic inflammation, in cultured supernatants were higher in PP and MPM patients compared with HD. Overall, asbestos-induced chronic inflammation in the lung as well as the pleural cavity may facilitate the onset of asbestos-induced cancers due to alterations in the interactions among fibers, immune cells such as T and B cells and macrophages, and mesothelial and lung epithelial cells. Further investigations regarding chronic inflammation caused by asbestos fibers may assist in identifying molecular targets for preventive and

  20. Mechanical properties of MDCK II cells exposed to gold nanorods

    Directory of Open Access Journals (Sweden)

    Anna Pietuch

    2015-01-01

    Full Text Available Background: The impact of gold nanoparticles on cell viability has been extensively studied in the past. Size, shape and surface functionalization including opsonization of gold particles ranging from a few nanometers to hundreds of nanometers are among the most crucial parameters that have been focussed on. Cytoxicity of nanomaterial has been assessed by common cytotoxicity assays targeting enzymatic activity such as LDH, MTT and ECIS. So far, however, less attention has been paid to the mechanical parameters of cells exposed to gold particles, which is an important reporter on the cellular response to external stimuli.Results: Mechanical properties of confluent MDCK II cells exposed to gold nanorods as a function of surface functionalization and concentration have been explored by atomic force microscopy and quartz crystal microbalance measurements in combination with fluorescence and dark-field microscopy.Conclusion: We found that cells exposed to CTAB coated gold nanorods display a concentration-dependent stiffening that cannot be explained by the presence of CTAB alone. The stiffening results presumably from endocytosis of particles removing excess membrane area from the cell’s surface. Another aspect could be the collapse of the plasma membrane on the actin cortex. Particles coated with PEG do not show a significant change in elastic properties. This observation is consistent with QCM measurements that show a considerable drop in frequency upon administration of CTAB coated rods suggesting an increase in acoustic load corresponding to a larger stiffness (storage modulus.

  1. ATM activation accompanies histone H2AX phosphorylation in A549 cells upon exposure to tobacco smoke

    Directory of Open Access Journals (Sweden)

    Traganos Frank

    2007-06-01

    Full Text Available Abstract Background In response to DNA damage or structural alterations of chromatin, histone H2AX may be phosphorylated on Ser139 by phosphoinositide 3-kinase related protein kinases (PIKKs such as ataxia telangiectasia mutated (ATM, ATM-and Rad-3 related (ATR kinase, or by DNA dependent protein kinase (DNA-PKcs. When DNA damage primarily involves formation of DNA double-strand breaks (DSBs, H2AX is preferentially phosphorylated by ATM rather than by the other PIKKs. We have recently reported that brief exposure of human pulmonary adenocarcinoma A549 cells or normal human bronchial epithelial cells (NHBE to cigarette smoke (CS induced phosphorylation of H2AX. Results We report here that H2AX phosphorylation in A549 cells induced by CS was accompanied by activation of ATM, as revealed by ATM phosphorylation on Ser1981 (ATM-S1981P detected immunocytochemically and by Western blotting. No cell cycle-phase specific differences in kinetics of ATM activation and H2AX phosphorylation were observed. When cells were exposed to CS from cigarettes with different tobacco and filter combinations, the expression levels of ATM-S1981P correlated well with the increase in expression of phosphorylated H2AX (γH2AX (R = 0.89. In addition, we note that while CS-induced γH2AX expression was localized within discrete foci, the activated ATM was distributed throughout the nucleoplasm. Conclusion These data implicate ATM as the PIKK that phosphorylates H2AX in response to DNA damage caused by CS. Based on current understanding of ATM activation, expression and localization, these data would suggest that, in addition to inducing potentially carcinogenic DSB lesions, CS may also trigger other types of DNA lesions and cause chromatin alterations. As checkpoint kinase (Chk 1, Chk2 and the p53 tumor suppressor gene are known to be phosphorylated by ATM, the present data indicate that exposure to CS may lead to their phosphorylation, with the downstream consequences

  2. Endothelial cell contraction increases Candida adherence to exposed extracellular matrix.

    Science.gov (United States)

    Klotz, S A; Maca, R D

    1988-01-01

    Bovine vascular endothelial cells treated with EDTA, urea, or thrombin underwent a marked, reversible contraction resulting in exposure of the subendothelial extracellular matrix (ECM). Candida yeasts adhered more to contracted monolayers than to confluent monolayers (P less than 0.01) by preferentially adhering to the ECM. Two strains of Candida albicans and one strain of Candida tropicalis bound avidly to exposed ECM, but Pseudomonas aeruginosa did not. However, treatment of endothelium with forskolin, which induces cell shape changes without exposure of the ECM, did not cause an increase in adherence. Images PMID:3137171

  3. The efficiency of photovoltaic cells exposed to pulsed laser light

    Science.gov (United States)

    Lowe, R. A.; Landis, G. A.; Jenkins, P.

    1993-01-01

    Future space missions may use laser power beaming systems with a free electron laser (FEL) to transmit light to a photovoltaic array receiver. To investigate the efficiency of solar cells with pulsed laser light, several types of GaAs, Si, CuInSe2, and GaSb cells were tested with the simulated pulse format of the induction and radio frequency (RF) FEL. The induction pulse format was simulated with an 800-watt average power copper vapor laser and the RF format with a frequency-doubled mode-locked Nd:YAG laser. Averaged current vs bias voltage measurements for each cell were taken at various optical power levels and the efficiency measured at the maximum power point. Experimental results show that the conversion efficiency for the cells tested is highly dependent on cell minority carrier lifetime, the width and frequency of the pulses, load impedance, and the average incident power. Three main effects were found to decrease the efficiency of solar cells exposed to simulated FEL illumination: cell series resistance, LC 'ringing', and output inductance. Improvements in efficiency were achieved by modifying the frequency response of the cell to match the spectral energy content of the laser pulse with external passive components.

  4. A short G1 phase imposes constitutive replication stress and fork remodelling in mouse embryonic stem cells

    DEFF Research Database (Denmark)

    Ahuja, Akshay K.; Jodkowska, Karolina; Teloni, Federico

    2016-01-01

    phosphorylation is dependent on Ataxia telangiectasia and Rad3 related (ATR) and is associated with chromatin loading of the ssDNA-binding proteins RPA and RAD51. Single-molecule analysis of replication intermediates reveals massive ssDNA gap accumulation, reduced fork speed and frequent fork reversal. All...... these marks of replication stress do not impair the mitotic process and are rapidly lost at differentiation onset. Delaying the G1/S transition in ESCs allows formation of 53BP1 nuclear bodies and suppresses ssDNA accumulation, fork slowing and reversal in the following S-phase. Genetic inactivation of fork...... slowing and reversal leads to chromosomal breakage in unperturbed ESCs. We propose that rapid cell cycle progression makes ESCs dependent on effective replication-coupled mechanisms to protect genome integrity....

  5. Mammalian cells exposed to ionizing radiation: structural and biochemical aspects

    Energy Technology Data Exchange (ETDEWEB)

    Sabanero, M.; Flores V, L. L. [Universidad de Guanajuato, Departamento de Biologia, DCNE, Noria Alta s/n, 36250 Guanajuato, Gto. (Mexico); Azorin V, J. C.; Vallejo, M. A.; Cordova F, T.; Sosa A, M. [Universidad de Guanajuato, Departamento de Ingenieria Fisica, DCI, Loma del Bosque 103, Col. Lomas del Campestre, 37150 Leon, Guanajuato (Mexico); Castruita D, J. P. [Universidad de Guadalajara, Departamento de Ecologia, CUCBA, Las Agujas, 45100 Zapopan, Jalisco (Mexico); Barbosa S, G., E-mail: myrna.sabanero@gmail.com [Universidad de Guanajuato, Departamento de Ciencias Medicas, DCS, 20 de Enero No. 929, Col. Obregon, 37000 Leon, Guanajuato (Mexico)

    2015-10-15

    Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv / year) and subsequently exposure to high doses have greater effects in people. However, it is unknown molecular and biochemical level alteration. This study, analyzes the susceptibility of a biological system (HeLa Atcc CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/ 90). Our evaluate multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin micro filaments), nuclei (D API), genomic DNA. The results indicate, that cells exposed to ionizing radiation structurally show alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin micro filaments. Similar alterations were observed in cells treated with a genotoxic agent (200μM H{sub 2}O{sub 2}/1 h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation. (Author)

  6. Cell-derived vesicles exposing coagulant tissue factor in saliva.

    Science.gov (United States)

    Berckmans, René J; Sturk, Auguste; van Tienen, Laurens M; Schaap, Marianne C L; Nieuwland, Rienk

    2011-03-17

    On vascular damage, coagulation is initiated by extravascular tissue factor (TF). Intravascular TF, which is present on circulating cell-derived vesicles, is noncoagulant under physiologic conditions but prothrombotic under pathologic conditions. Human saliva triggers coagulation, but the mechanism and physiologic relevance are unknown. Because saliva is known to contain TF, we hypothesized that this TF may also be associated with cell-derived vesicles to facilitate coagulation when saliva directly contacts blood. The saliva-induced shortening of the clotting time of autologous plasma and whole blood from healthy subjects (n = 10) proved TF-dependent. This TF was associated with various types of cell-derived vesicles, including microparticles and exosomes. The physiologic function was shown by adding saliva to human pericardial wound blood collected from patients undergoing cardiac surgery. Addition of saliva shortened the clotting time from 300 ± 96 to 186 ± 24 seconds (P = .03). Our results show that saliva triggers coagulation, thereby reducing blood loss and the risk of pathogens entering the blood. We postulate that our reflex to lick a wound may be a mechanism to enable TF-exposing vesicles, present in saliva, to aid in the coagulation process and thus protect the organism from entering pathogens. This unique compartmentalization may be highly conserved because also animals lick their wounds.

  7. Chronic Replication Problems Impact Cell Morphology and Adhesion of DNA Ligase I Defective Cells.

    Directory of Open Access Journals (Sweden)

    Paolo Cremaschi

    Full Text Available Moderate DNA damage resulting from metabolic activities or sub-lethal doses of exogenous insults may eventually lead to cancer onset. Human 46BR.1G1 cells bear a mutation in replicative DNA ligase I (LigI which results in low levels of replication-dependent DNA damage. This replication stress elicits a constitutive phosphorylation of the ataxia telangiectasia mutated (ATM checkpoint kinase that fails to arrest cell cycle progression or to activate apoptosis or cell senescence. Stable transfection of wild type LigI, as in 7A3 cells, prevents DNA damage and ATM activation. Here we show that parental 46BR.1G1 and 7A3 cells differ in important features such as cell morphology, adhesion and migration. Comparison of gene expression profiles in the two cell lines detects Bio-Functional categories consistent with the morphological and migration properties of LigI deficient cells. Interestingly, ATM inhibition makes 46BR.1G1 more similar to 7A3 cells for what concerns morphology, adhesion and expression of cell-cell adhesion receptors. These observations extend the influence of the DNA damage response checkpoint pathways and unveil a role for ATM kinase activity in modulating cell biology parameters relevant to cancer progression.

  8. MEK inhibitors block growth of lung tumours with mutations in ataxia-telangiectasia mutated

    Czech Academy of Sciences Publication Activity Database

    Smida, M.; de la Cruz, F.F.; Kerzendorfer, C.; Uras, I.Z.; Mair, B.; Mazouzi, A.; Suchánková, Tereza; Konopka, T.; Katz, A.M.; Paz, K.; Nagy-Bojarszky, K.; Muellner, M.K.; Bago-Horvath, Z.; Haura, E.B.; Loizou, J.I.; Nijman, S.M.B.

    2016-01-01

    Roč. 7, DEC2016 (2016), č. článku 13701. ISSN 2041-1723 Institutional support: RVO:68081707 Keywords : breast- cancer * insulin-resistance * missense mutations Subject RIV: BO - Biophysics Impact factor: 12.124, year: 2016

  9. Ataxia telangiectasia - A report of a case in Port Harcourt | Yaguo ...

    African Journals Online (AJOL)

    She achieved normal early developmental milestones. She is the second child of adoptive parents and was adopted at 2weeks of age. Her biological mother was said to have died immediately after delivery. Her adoptive parents were of high socioeconomic class. The detail of child's family history was not known to the ...

  10. T-cell-specific deletion of Mof blocks their differentiation and results in genomic instability in mice.

    Science.gov (United States)

    Gupta, Arun; Hunt, Clayton R; Pandita, Raj K; Pae, Juhee; Komal, K; Singh, Mayank; Shay, Jerry W; Kumar, Rakesh; Ariizumi, Kiyoshi; Horikoshi, Nobuo; Hittelman, Walter N; Guha, Chandan; Ludwig, Thomas; Pandita, Tej K

    2013-05-01

    Ataxia telangiectasia patients develop lymphoid malignancies of both B- and T-cell origin. Similarly, ataxia telangiectasia mutated (Atm)-deficient mice exhibit severe defects in T-cell maturation and eventually develop thymomas. The function of ATM is known to be influenced by the mammalian orthologue of the Drosophila MOF (males absent on the first) gene. Here, we report the effect of T-cell-specific ablation of the mouse Mof (Mof) gene on leucocyte trafficking and survival. Conditional Mof(Flox/Flox) (Mof (F/F)) mice expressing Cre recombinase under control of the T-cell-specific Lck proximal promoter (Mof(F/F)/Lck-Cre(+)) display a marked reduction in thymus size compared with Mof(F/F)/Lck-Cre(-) mice. In contrast, the spleen size of Mof(F/F)/Lck-Cre(+) mice was increased compared with control Mof(F/F)/Lck-Cre(-) mice. The thymus of Mof(F/F)/Lck-Cre(+) mice contained significantly reduced T cells, whereas thymic B cells were elevated. Within the T-cell population, CD4(+)CD8(+) double-positive T-cell levels were reduced, whereas the immature CD4(-)CD8(-) double-negative (DN) population was elevated. Defective T-cell differentiation is also evident as an increased DN3 (CD44(-)CD25(+)) population, the cell stage during which T-cell receptor rearrangement takes place. The differentiation defect in T cells and reduced thymus size were not rescued in a p53-deficient background. Splenic B-cell distributions were similar between Mof(F/F)/Lck-Cre(+) and Mof(F/F)/Lck-Cre(-) mice except for an elevation of the κ light-chain population, suggestive of an abnormal clonal expansion. T cells from Mof(F/F)/Lck-Cre(+) mice did not respond to phytohaemagglutinin (PHA) stimulation, whereas LPS-stimulated B cells from Mof(F/F)/Lck-Cre(+) mice demonstrated spontaneous genomic instability. Mice with T-cell-specific loss of MOF had shorter lifespans and decreased survival following irradiation than did Mof(F/F)/Lck-Cre(-) mice. These observations suggest that Mof plays a critical

  11. Differential responses to high- and low-dose ultraviolet-B stress in tobacco Bright Yellow-2 cells

    Directory of Open Access Journals (Sweden)

    Shinya eTakahashi

    2015-04-01

    Full Text Available Ultraviolet (UV-B irradiation leads to DNA damage, cell cycle arrest, growth inhibition, and cell death. To evaluate the UV-B stress–induced changes in plant cells, we developed a model system based on tobacco Bright Yellow-2 (BY-2 cells. Both low-dose UV-B (low UV-B: 740 J m−2 and high-dose UV-B (high UV-B: 2960 J m−2 inhibited cell proliferation and induced cell death; these effects were more pronounced at high UV-B. Flow cytometry showed cell cycle arrest within 1 day after UV-B irradiation; neither low- nor high-UV-B–irradiated cells entered mitosis within 12 h. Cell cycle progression was gradually restored in low-UV-B–irradiated cells but not in high-UV-B–irradiated cells. UV-A irradiation, which activates cyclobutane pyrimidine dimer (CPD photolyase, reduced inhibition of cell proliferation by low but not high UV-B and suppressed high-UV-B–induced cell death. UV-B induced CPD formation in a dose-dependent manner. The amounts of CPDs decreased gradually within 3 days in low-UV-B–irradiated cells, but remained elevated after 3 days in high-UV-B–irradiated cells. Low UV-B slightly increased the number of DNA single-strand breaks detected by the comet assay at 1 day after irradiation, and then decreased at 2 and 3 days after irradiation. High UV-B increased DNA fragmentation detected by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay 1 and 3 days after irradiation. Caffeine, an inhibitor of ataxia telangiectasia mutated (ATM and ataxia telangiectasia and Rad3-related (ATR checkpoint kinases, reduced the rate of cell death in high-UV-B–irradiated cells. Our data suggest that low-UV-B–induced CPDs and/or DNA strand-breaks inhibit DNA replication and proliferation of BY-2 cells, whereas larger contents of high-UV-B–induced CPDs and/or DNA strand-breaks lead to cell death.

  12. Activation of ATM by DNA Damaging Agents

    National Research Council Canada - National Science Library

    Kurz, Ebba U; Lees-Miller, Susan P

    2005-01-01

    Ataxia-telangiectasia mutated (ATM) is a serine/threonine protein kinase that acts as a master switch controlling the cell cycle in response to ionizing radiation-induced DNA double-strand breaks (DSBs...

  13. Activation of ATM by DNA Damaging Agents

    National Research Council Canada - National Science Library

    Kurz, Ebba U; Lees-Miller, Susan P

    2004-01-01

    Ataxia-telangiectasia mutated (ATM) is a serine/threonine protein kinase that acts as a master switch controlling the cell cycle in response to ionizing radiation-induced DNA double-strand breaks (DSBs...

  14. The Saccharomyces cerevisiae MEC1 gene, which encodes a homolog of the human ATM gene product, is required for G1 arrest following radiation treatment.

    OpenAIRE

    Siede, W.; Allen, J B; Elledge, S. J.; Friedberg, E C

    1996-01-01

    The Saccharomyces cerevisiae gene MEC1 represents a structural homolog of the human gene ATM mutated in ataxia telangiectasia patients. Like human ataxia telangiectasia cell lines, mec1 mutants are defective in G2 and S-phase cell cycle checkpoints in response to radiation treatment. Here we show an additional defect in G1 arrest following treatment with UV light or gamma rays and map a defective arrest stage at or upstream of START in the yeast cell cycle.

  15. Improved immunogenicity of fusions between ethanol-treated cancer cells and dendritic cells exposed to dual TLR stimulation

    National Research Council Canada - National Science Library

    Koido, Shigeo; Homma, Sadamu; Okamoto, Masato; Namiki, Yoshihisa; Kan, Shin; Takakura, Kazuki; Kajihara, Mikio; Uchiyama, Kan; Hara, Eiich; Ohkusa, Toshifumi; Gong, Jianlin; Tajiri, Hisao

    2013-01-01

    ...) fused to whole cancer cells. We have recently revealed that ethanol-treated neoplastic cells fused to DCs exposed to 2 Toll-like receptor agonists efficiently induce cytotoxic T lymphocytes via TGF...

  16. B cells exposed to enterobacterial components suppress development of experimental colitis

    DEFF Research Database (Denmark)

    Schmidt, Esben Gjerløff Wedebye; Larsen, Hjalte List; Kristensen, Nanna Ny

    2012-01-01

    ). RESULTS: We demonstrate that splenic B cells exposed to ebx produce large amounts of IL-10 in vitro and express CD1d and CD5 previously known to be associated with regulatory B cells. In SCID mice transplanted with colitogenic CD4(+) CD25(-) T cells, co-transfer of ebx-B cells significantly suppressed...... development of colitis. Suppression was dependent on B cell-derived IL-10, as co-transfer of IL-10 knockout ebx-B cells failed to suppress colitis. Ebx-B cell-mediated suppression of colitis was associated with a decrease in interferon gamma (IFN-¿)-producing T(H) 1 cells and increased frequencies of Foxp3......-expressing T cells. CONCLUSIONS: These data demonstrate that splenic B cells exposed to enterobacterial components acquire immunosuppressive functions by which they can suppress development of experimental T cell-mediated colitis in an IL-10-dependent way. (Inflamm Bowel Dis 2011;)....

  17. Helichrysetin Induces DNA Damage that Triggers JNK-Mediated Apoptosis in Ca Ski Cells.

    Science.gov (United States)

    Fong, Ho Yen; Abd Malek, Sri Nurestri; Yee, Hui Shin; Karsani, Saiful Anuar

    2017-01-01

    Cervical cancer has become one of the most common cancers in women and currently available treatment options for cervical cancer are very limited. Naturally occurring chalcones and its derivatives have been studied extensively as a potential anticancer agent in different types of cancer and helichrysetin is naturally occurring chalcone that possess potent antiproliferative activity toward human cancer cells. Inhibitory activity of helichrysetin was evaluated at different concentrations. Ability of helichrysetin to induce apoptosis and its relation with c-Jun N-terminal kinase (JNK)-mediated mechanism of apoptosis was assessed using flow cytometry and Western blotting. Helichrysetin inhibited Ca Ski cells at half maximal inhibitory concentration 30.62 ± 0.38 μM. This compound has the ability to induce DNA damage, mitochondrial membrane disruption, and loss of cell membrane integrity. We have shown that apoptosis was induced through the activation of JNK-mediated apoptosis by DNA damage in the cells then triggering p53-downstream apoptotic pathway with increased expression of pro-apoptotic proteins, Bax and caspase 3, and suppression of Bcl-2 anti-apoptotic protein. DNA damage in the cells also caused phosphorylation of protein ataxia-telangiectasia mutated, an activator of DNA damage response. We conclude that helichrysetin can inhibit Ca Ski cells through DNA damage-induced JNK-mediated apoptotic pathway highlighting the potential of this compound as anticancer agent for cervical cancer. Helichrysetin induced DNA damage in Ca Ski cellsDNA damage caused JNK-mediated phosphorylation of p53 resulting in p53-mediated apoptosisHelichrysetin is a potential DNA damage inducing agent through JNK activation to kill human cervical carcinoma cells. Abbreviations used: ATM: Ataxia-telangiectasia mutated, DAPI: 4',6-diamidino-2-phenylindole, DMSO: Dimethyl sulfoxide, FITC: Fluorescein isothiocyanate, IC 50 : Half maximal inhibitory concentration, JC1-5,5',6,6'-Tetrachloro: 1

  18. Mitochondrial hyperpolarization and cytochrome-c release in microwave-exposed MCF-7 cells.

    Science.gov (United States)

    Esmekaya, Meric A; Canseven, Ayşe G; Kayhan, Handan; Tuysuz, Mehmet Z; Sirav, Bahriye; Seyhan, Nesrin

    2017-04-01

    This study examines the effects of a 2.1-GHz WCDMA-modulated microwave (MW) radiation on apoptotic activity and mitochondrial membrane potential (ΔΨm) in MCF-7 cells. The cells were exposed to the MW at a specific absorption rate (SAR) of 0.528 W/kg for 4 or 24 h. The antiproliferative effect of MW exposure was determined by the MTT test. Cytochrome-c and p53 levels were determined by an ELISA method. The relative ΔΨm was analysed by JC-1 staining using flow cytometer. Apoptotic rate of the cells was measured by Annexin-V-FITC staining. All assays were performed after certain time of incubations (15 min-4 h) following MW exposure. MW-exposed cells showed a significant decrease in viability when compared to unexposed cells. A significantly larger decrease was observed after longer exposure. The percentage of apoptotic cells, amount of cytochrome-c, and relative ΔΨm were significantly higher in MW-exposed cells. The percent of apoptotic cells and relative ΔΨm in 24 h MW-exposed group was significantly higher than those in 4 h MW-exposed group. However, no significant change was observed in p53 levels. These results demonstrated that exposure to 2.1-GHz WCDMA-modulated MW radiation caused hyperpolarization of mitochondria that in turn induced apoptosis in MCF-7 cells.

  19. Radiotherapy induces cell cycle arrest and cell apoptosis in nasopharyngeal carcinoma via the ATM and Smad pathways.

    Science.gov (United States)

    Li, Ming-Yi; Liu, Jin-Quan; Chen, Dong-Ping; Li, Zhou-Yu; Qi, Bin; He, Lu; Yu, Yi; Yin, Wen-Jin; Wang, Meng-Yao; Lin, Ling

    2017-09-02

    Nasopharyngeal carcinoma (NPC) is a common malignant neoplasm of the head and neck which is harmful to human's health. Radiotherapy is commonly used in the treatment of NPC and it induces immediate cell cycle arrest and cell apoptosis. However, the mechanism remains unknown. Evidences suggested the activation of Ataxia telangiectasia mutated (ATM) pathway and Smad pathway are 2 of the important crucial mediators in the function of radiotherapy. In this study, we performed in vitro assays with human nasopharyngeal carcinoma CNE-2 cells and in vivo assays with nude mice to investigate the role of the ATM and Smad pathways in the treatment of nasopharyngeal carcinoma with radiotherapy. The results suggested that radiation induced activation of ATM pathway by inducing expression of p-ATM, p-CHK1, p-CHK2, p15 and inhibiting expression of p-Smad3. In addition, Caspase3 expression was increased while CDC25A was decreased, leading to cell cycle arrest and cell apoptosis. On the other hand, activation of Smad3 can inhibited the ATM pathway and attenuated the efficacy of radiation. In summary, we suggest that both ATM and Smad pathways contribute to the cell cycle arrest and cell apoptosis during nasopharyngeal carcinoma cells treated with radiation.

  20. Targeting Ongoing DNA Damage in Multiple Myeloma: Effects of DNA Damage Response Inhibitors on Plasma Cell Survival

    Directory of Open Access Journals (Sweden)

    Ana Belén Herrero

    2017-05-01

    Full Text Available Human myeloma cell lines (HMCLs and a subset of myeloma patients with poor prognosis exhibit high levels of replication stress (RS, leading to DNA damage. In this study, we confirmed the presence of DNA double-strand breaks (DSBs in several HMCLs by measuring γH2AX and RAD51 foci and analyzed the effect of various inhibitors of the DNA damage response on MM cell survival. Inhibition of ataxia telangiectasia and Rad3-related protein (ATR, the main kinase mediating the response to RS, using the specific inhibitor VE-821 induced more cell death in HMCLs than in control lymphoblastoid cells and U266, an HMCL with a low level of DNA damage. The absence of ATR was partially compensated by ataxia telangiectasia-mutated protein (ATM, since chemical inhibition of both kinases using VE-821 and KU-55933 significantly increased the death of MM cells with DNA damage. We found that ATM and ATR are involved in DSB repair by homologous recombination (HR in MM. Inhibition of both kinases resulted in a stronger inhibition that may underlie cell death induction, since abolition of HR using two different inhibitors severely reduced survival of HMCLs that exhibit DNA damage. On the other hand, inhibition of the other route involved in DSB repair, non-homologous end joining (NHEJ, using the DNA-PK inhibitor NU7441, did not affect MM cell viability. Interestingly, we found that NHEJ inhibition did not increase cell death when HR was simultaneously inhibited with the RAD51 inhibitor B02, but it clearly increased the level of cell death when HR was inhibited with the MRE11 inhibitor mirin, which interferes with recombination before DNA resection takes place. Taken together, our results demonstrate for the first time that MM cells with ongoing DNA damage rely on an intact HR pathway, which thereby suggests therapeutic opportunities. We also show that inhibition of HR after the initial step of end resection might be more appropriate for inducing MM cell death, since it

  1. Targeting Ongoing DNA Damage in Multiple Myeloma: Effects of DNA Damage Response Inhibitors on Plasma Cell Survival.

    Science.gov (United States)

    Herrero, Ana Belén; Gutiérrez, Norma Carmen

    2017-01-01

    Human myeloma cell lines (HMCLs) and a subset of myeloma patients with poor prognosis exhibit high levels of replication stress (RS), leading to DNA damage. In this study, we confirmed the presence of DNA double-strand breaks (DSBs) in several HMCLs by measuring γH2AX and RAD51 foci and analyzed the effect of various inhibitors of the DNA damage response on MM cell survival. Inhibition of ataxia telangiectasia and Rad3-related protein (ATR), the main kinase mediating the response to RS, using the specific inhibitor VE-821 induced more cell death in HMCLs than in control lymphoblastoid cells and U266, an HMCL with a low level of DNA damage. The absence of ATR was partially compensated by ataxia telangiectasia-mutated protein (ATM), since chemical inhibition of both kinases using VE-821 and KU-55933 significantly increased the death of MM cells with DNA damage. We found that ATM and ATR are involved in DSB repair by homologous recombination (HR) in MM. Inhibition of both kinases resulted in a stronger inhibition that may underlie cell death induction, since abolition of HR using two different inhibitors severely reduced survival of HMCLs that exhibit DNA damage. On the other hand, inhibition of the other route involved in DSB repair, non-homologous end joining (NHEJ), using the DNA-PK inhibitor NU7441, did not affect MM cell viability. Interestingly, we found that NHEJ inhibition did not increase cell death when HR was simultaneously inhibited with the RAD51 inhibitor B02, but it clearly increased the level of cell death when HR was inhibited with the MRE11 inhibitor mirin, which interferes with recombination before DNA resection takes place. Taken together, our results demonstrate for the first time that MM cells with ongoing DNA damage rely on an intact HR pathway, which thereby suggests therapeutic opportunities. We also show that inhibition of HR after the initial step of end resection might be more appropriate for inducing MM cell death, since it prevents the

  2. Cell-derived vesicles exposing coagulant tissue factor in saliva

    NARCIS (Netherlands)

    Berckmans, René J.; Sturk, Auguste; van Tienen, Laurens M.; Schaap, Marianne C. L.; Nieuwland, Rienk

    2011-01-01

    On vascular damage, coagulation is initiated by extravascular tissue factor (TF). Intravascular TF, which is present on circulating cell-derived vesicles, is non-coagulant under physiologic conditions but prothrombotic under pathologic conditions. Human saliva triggers coagulation, but the mechanism

  3. Correlation between dielectric property by dielectrophoretic levitation and growth activity of cells exposed to electric field.

    Science.gov (United States)

    Hakoda, Masaru; Hirota, Yusuke

    2013-09-01

    The purpose of this study is to develop a system analyzing cell activity by the dielectrophoresis method. Our previous studies revealed a correlation between the growth activity and dielectric property (Re[K(ω)]) of mouse hybridoma 3-2H3 cells using dielectrophoretic levitation. Furthermore, it was clarified that the differentiation activity of many stem cells could be evaluated by the Re[K(ω)] without differentiation induction. In this paper, 3-2H3 cells exposed to an alternating current (AC) electric field or a direct current (DC) electric field were cultivated, and the influence of damage by the electric field on the growth activity of the cells was examined. To evaluate the activity of the cells by measuring the Re[K(ω)], the correlation between the growth activity and the Re[K(ω)] of the cells exposed to the electric field was examined. The relations between the cell viability, growth activity, and Re[K(ω)] in the cells exposed to the AC electric field were obtained. The growth activity of the cells exposed to the AC electric field could be evaluated by the Re[K(ω)]. Furthermore, it was found that the adverse effects of the electric field on the cell viability and the growth activity were smaller in the AC electric field than the DC electric field.

  4. Caffeine markedly sensitizes human mesothelioma cell lines to pemetrexed

    Science.gov (United States)

    Min, Sang Hee; Goldman, I. David; Zhao, Rongbao

    2013-01-01

    Pemetrexed is a new generation antifolate approved for the treatment of mesothelioma and non-small cell lung cancer. Caffeine is known to augment radiation or chemotherapeutic drug-induced cell killing. The current study addresses the impact of caffeine on the activity of pemetrexed in mesothelioma cell lines. Caffeine enhanced pemetrexed activity in all four mesothelioma cell lines tested (H2052, H2373, H28 and MSTO-211H). Caffeine sensitized H2052 cells in a dose- and schedule-dependent manner, and was associated with a markedly decreased clonogenic survival. Caffeine sensitization occurred only in cells subjected to pulse, but not continuous, exposure to pemetrexed. Similar pemetrexed sensitization was also observed with the clinically better tolerated caffeine analog, theobromine. Pemetrexed sensitization by caffeine was associated with an increase in pemetrexed-induced phosphorylation of ataxia-telangiectasia-mutated (ATM) and Chk1. These data indicate that caffeine and its analog, theobromine, may be a useful approach to enhance pemetrexed-based chemotherapy. PMID:17594092

  5. Autosomal mutants of proton-exposed kidney cells display frequent loss of heterozygosity on nonselected chromosomes.

    Science.gov (United States)

    Grygoryev, Dmytro; Dan, Cristian; Gauny, Stacey; Eckelmann, Bradley; Ohlrich, Anna P; Connolly, Marissa; Lasarev, Michael; Grossi, Gianfranco; Kronenberg, Amy; Turker, Mitchell S

    2014-05-01

    High-energy protons found in the space environment can induce mutations and cancer, which are inextricably linked. We hypothesized that some mutants isolated from proton-exposed kidneys arose through a genome-wide incident that causes loss of heterozygosity (LOH)-generating mutations on multiple chromosomes (termed here genomic LOH). To test this hypothesis, we examined 11 pairs of nonselected chromosomes for LOH events in mutant cells isolated from the kidneys of mice exposed to 4 or 5 Gy of 1 GeV protons. The mutant kidney cells were selected for loss of expression of the chromosome 8-encoded Aprt gene. Genomic LOH events were also assessed in Aprt mutants isolated from isogenic cultured kidney epithelial cells exposed to 5 Gy of protons in vitro. Control groups were spontaneous Aprt mutants and clones isolated without selection from the proton-exposed kidneys or cultures. The in vivo results showed significant increases in genomic LOH events in the Aprt mutants from proton-exposed kidneys when compared with spontaneous Aprt mutants and when compared with nonmutant (i.e., nonselected) clones from the proton-exposed kidneys. A bias for LOH events affecting chromosome 14 was observed in the proton-induced Aprt mutants, though LOH for this chromosome did not confer increased radiation resistance. Genomic LOH events were observed in Aprt mutants isolated from proton-exposed cultured kidney cells; however the incidence was fivefold lower than in Aprt mutants isolated from exposed intact kidneys, suggesting a more permissive environment in the intact organ and/or the evolution of kidney clones prior to their isolation from the tissue. We conclude that proton exposure creates a subset of viable cells with LOH events on multiple chromosomes, that these cells form and persist in vivo, and that they can be isolated from an intact tissue by selection for a mutation on a single chromosome.

  6. DNA Fragmentation in mammalian cells exposed to various light ions

    Science.gov (United States)

    Belli, M.; Cherubini, R.; Dalla Vecchia, M.; Dini, V.; Esposito, G.; Moschini, G.; Sapora, O.; Signoretti, C.; Simone, G.; Sorrentino, E.; Tabocchini, M. A.

    Elucidation of how effects of densely ionizing radiation at cellular level are linked to DNA damage is fundamental for a better understanding of the mechanisms leading to genomic damage (especially chromosome aberrations) and developing biophysical models to predict space radiation effects. We have investigated the DNA fragmentation patterns induced in Chinese hamster V79 cells by 31 keV/μm protons, 123 keV/μm helium-4 ions and γ-rays in the size range 0.023-5.7 Mbp, using calibrated Pulsed Field Gel Electrophoresis (PFGE). The frequency distributions of fragments induced by the charged particles were shifted towards smaller sizes with respct to that induced by comparable doses of γ-rays. The DSB yields, evaluated from the fragments induced in the size range studied, were higher for protons and helium ions than for γ-rays by a factor of about 1.9 and 1.2, respectively. However, these ratios do not adequately reflect the RBE observed on the same cells for inactivation and mutation induced by these beams. This is a further indication for the lack of correlation between the effects exerted at cellular level and the initial yield of DSB. The dependence on radiation quality of the fragmentation pattern suggests that it may have a role in damage reparability. We have analyzed these patterns with a "random breakage" model generalized in order to consider the initial non-random distribution of the DNA molecules. Our results suggest that a random breakage mechanism can describe with a reasonable approximation the DNA fragmentation induced by γ-rays, while the approximation is not so good for light ions, likely due to the interplay between ion tracks and chromatin organization at the loop level.

  7. JS-K, a nitric oxide prodrug, induces DNA damage and apoptosis in HBV-positive hepatocellular carcinoma HepG2.2.15 cell.

    Science.gov (United States)

    Liu, Zhengyun; Li, Guangmin; Gou, Ying; Xiao, Dongyan; Luo, Guo; Saavedra, Joseph E; Liu, Jie; Wang, Huan

    2017-08-01

    Hepatocellular carcinoma (HCC) is the most important cause of cancer-related death, and 85% of HCC is caused by chronic HBV infection, the prognosis of patients and the reduction of HBV DNA levels remain unsatisfactory. JS-K, a nitric oxide-releasing diazeniumdiolates, is effective against various tumors, but little is known on its effects on HBV positive HCC. We found that JS-K reduced the expression of HBsAg and HBeAg in HBV-positive HepG2.2.15 cells. This study aimed to further examine anti-tumor effects of JS-K on HepG2.2.15 cells. The MTT assay and colony forming assay were used to study the cell growth inhibition of JS-K; scratch assay and transwell assay were performed to detect cell migration. The cell cycle was detected by flow cytometry. The immunofluorescence, flow cytometry analysis, and western blot were used to study DNA damage and cell apoptosis. JS-K inhibited HepG2.2.15 cell growth in a dose-dependent manner, suppressed cell colony formation and migration, arrested cells gather in the G2 phase. JS-K (1-20μM) increased the expression of DNA damage-associated protein phosphorylation H2AX (γH2AX), phosphorylation of checkpoint kinase 1 (p-Chk1), phosphorylation of checkpoint kinase 2 (p-Chk2), ataxia-telangiectasia mutated (ATM), phosphorylation of ataxia-telangiectasia mutated rad3-related (p-ATR) and apoptotic-associated proteins cleaved caspase-3, cleaved caspase-7, cleaved poly ADP-ribose polymerase (cleaved PARP). The study demonstrated JS-K is effective against HBV-positive HepG2.2.15 cells, the mechanisms are not only related to inhibition of HBsAg and HBeAg secretion, but also related with induction of DNA damage and apoptosis. JS-K is a promising anti-cancer candidate against HBV-positive HCC. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  8. Analysis of Chromatin Opening in Heterochromatic Non-Small Cell Lung Cancer Tumor-Initiating Cells in Relation to DNA-Damaging Antitumor Treatment.

    Science.gov (United States)

    Eriksson, Mina; Hååg, Petra; Brzozowska, Beata; Lipka, Magdalena; Lisowska, Halina; Lewensohn, Rolf; Wojcik, Andrzej; Viktorsson, Kristina; Lundholm, Lovisa

    2018-01-01

    We previously reported that sphere-forming non-small cell lung cancer (NSCLC) tumor-initiating cells (TICs) have an altered activation of DNA damage response- and repair proteins and are refractory to DNA-damaging treatments. We analyzed whether chromatin organization plays a role in the observed refractoriness. Bulk cells and TICs from the NSCLC H23 and H1299 cell lines were examined using cell viability, clonogenic survival, Western blot, short interfering RNA analysis, and micronucleus assay. NSCLC TICs displayed elevated heterochromatin markers trimethylated lysine 9 of histone H3 and heterochromatin protein 1γ relative to bulk cells and reduced cell viability upon histone deacetylase inhibition (HDACi). Vorinostat and trichostatin A increased the euchromatin markers acetylated lysine 9/14 of histone H3 and lysine 8 of histone H4, and HDACi pretreatment increased the phosphorylation of the DNA damage response proteins ataxia telangiectasia mutated and DNA-dependent protein kinase, catalytic subunit, upon irradiation in TICs. HDACi sensitized TICs to cisplatin and to some extent to ionizing irradiation. The protectiveness of a dense chromatin structure was indicated by an enhanced frequency of micronuclei in TICs following irradiation, after knockdown of heterochromatin protein 1γ. Although confirmatory studies in additional NSCLC model systems and with respect to analyses of other DNA damage response proteins are needed, our data point toward a heterochromatic structure of NSCLC TICs, such that HDACi can sensitize TICs to DNA damage. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Cancer cells with alternative lengthening of telomeres do not display a general hypersensitivity to ATR inhibition

    Directory of Open Access Journals (Sweden)

    Katharina I Deeg

    2016-08-01

    Full Text Available Telomere maintenance is a hallmark of cancer as it provides cancer cells with cellular immortality. A significant fraction of tumors uses the alternative lengthening of telomeres (ALT pathway to elongate their telomeres and to gain an unlimited proliferation potential. Since the ALT pathway is unique to cancer cells, it represents a potentially valuable, currently unexploited target for anticancer therapies. Recently, it was proposed that ALT renders cells hypersensitive to ataxia telangiectasia- and RAD3-related (ATR protein inhibitors (Flynn et al., Science 347, 273. Here, we measured the response of various ALT or telomerase positive cell lines to the ATR inhibitor VE-821. In addition, we compared the effect of the inhibitor on cell viability in isogenic cell lines, in which ALT was active or suppressed. In these experiments a general ATR inhibitor sensitivity of cells with ALT could not be confirmed. We rather propose that the observed variations in sensitivity reflect differences between cell lines that are unrelated to ALT.

  10. Dose rate effect on low-dose hyper-radiosensitivity with cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Geon-Min; Kim, Eun-Hee [Seoul National University, Seoul (Korea, Republic of)

    2016-10-15

    Low-dose hyper-radiosensitivity (HRS) is the phenomenon that mammalian cells exhibit higher sensitivity to radiation at low doses (< 0.5 Gy) than expected by the linear-quadratic model. At doses above 0.5Gy, the cellular response is recovered to the level expected by the linear-quadratic model. This transition is called the increased radio-resistance (IRR). HRS was first verified using Chinese hamster V79 cells in vitro by Marples and has been confirmed in studies with other cell lines including human normal and tumor cells. HRS is known to be induced by inactivation of ataxia telangiectasia-mutated (ATM), which plays a key role in repairing DNA damages. Considering the connection between ATM and HRS, one can infer that dose rate may affect cellular response regarding HRS at low doses. In this study, we quantitated the effect of dose rate on HRS by clonogenic assay with normal and tumor cells. The HRS of cells at low dose exposures is a phenomenon already known. In this study, we observed HRS of rat normal diencephalon cells and rat gliosarcoma cells at doses below 1 Gy. In addition, we found that dose rate mattered. HRS occurred at low doses, but only when total dose was delivered at a rate below certain level.

  11. Estrogen enhanced cell-cell signalling in breast cancer cells exposed to targeted irradiation

    Directory of Open Access Journals (Sweden)

    Held Kathryn D

    2008-06-01

    Full Text Available Abstract Background Radiation-induced bystander responses, where cells respond to their neighbours being irradiated are being extensively studied. Although evidence shows that bystander responses can be induced in many types of cells, it is not known whether there is a radiation-induced bystander effect in breast cancer cells, where the radiosensitivity may be dependent on the role of the cellular estrogen receptor (ER. This study investigated radiation-induced bystander responses in estrogen receptor-positive MCF-7 and estrogen receptor-negative MDA-MB-231 breast cancer cells. Methods The influence of estrogen and anti-estrogen treatments on the bystander response was determined by individually irradiating a fraction of cells within the population with a precise number of helium-3 using a charged particle microbeam. Damage was scored as chromosomal damage measured as micronucleus formation. Results A bystander response measured as increased yield of micronucleated cells was triggered in both MCF-7 and MDA-MB-231 cells. The contribution of the bystander response to total cell damage in MCF-7 cells was higher than that in MDA-MB-231 cells although the radiosensitivity of MDA-MB-231 was higher than MCF-7. Treatment of cells with 17β-estradiol (E2 increased the radiosensitivity and the bystander response in MCF-7 cells, and the effect was diminished by anti-estrogen tamoxifen (TAM. E2 also increased the level of intracellular reactive oxygen species (ROS in MCF-7 cells in the absence of radiation. In contrast, E2 and TAM had no influence on the bystander response and ROS levels in MDA-MB-231 cells. Moreover, the treatment of MCF-7 cells with antioxidants eliminated both the E2-induced ROS increase and E2-enhanced bystander response triggered by the microbeam irradiation, which indicates that ROS are involved in the E2-enhanced bystander micronuclei formation after microbeam irradiation. Conclusion The observation of bystander responses in breast

  12. ATR Kinase Inhibition Protects Non-cycling Cells from the Lethal Effects of DNA Damage and Transcription Stress*

    Science.gov (United States)

    Kemp, Michael G.; Sancar, Aziz

    2016-01-01

    ATR (ataxia telangiectasia and Rad-3-related) is a protein kinase that maintains genome stability and halts cell cycle phase transitions in response to DNA lesions that block DNA polymerase movement. These DNA replication-associated features of ATR function have led to the emergence of ATR kinase inhibitors as potential adjuvants for DNA-damaging cancer chemotherapeutics. However, whether ATR affects the genotoxic stress response in non-replicating, non-cycling cells is currently unknown. We therefore used chemical inhibition of ATR kinase activity to examine the role of ATR in quiescent human cells. Although ATR inhibition had no obvious effects on the viability of non-cycling cells, inhibition of ATR partially protected non-replicating cells from the lethal effects of UV and UV mimetics. Analyses of various DNA damage response signaling pathways demonstrated that ATR inhibition reduced the activation of apoptotic signaling by these agents in non-cycling cells. The pro-apoptosis/cell death function of ATR is likely due to transcription stress because the lethal effects of compounds that block RNA polymerase movement were reduced in the presence of an ATR inhibitor. These results therefore suggest that whereas DNA polymerase stalling at DNA lesions activates ATR to protect cell viability and prevent apoptosis, the stalling of RNA polymerases instead activates ATR to induce an apoptotic form of cell death in non-cycling cells. These results have important implications regarding the use of ATR inhibitors in cancer chemotherapy regimens. PMID:26940878

  13. Metformin is also effective on lactic acidosis-exposed melanoma cells switched to oxidative phosphorylation.

    Science.gov (United States)

    Peppicelli, Silvia; Toti, Alessandra; Giannoni, Elisa; Bianchini, Francesca; Margheri, Francesca; Del Rosso, Mario; Calorini, Lido

    2016-07-17

    Low extracellular pH promotes in melanoma cells a malignant phenotype characterized by an epithelial-to-mesenchymal transition (EMT) program, endowed with mesenchymal markers, high invasiveness and pro-metastatic property. Here, we demonstrate that melanoma cells exposed to an acidic extracellular microenvironment, 6.7±0.1, shift to an oxidative phosphorylation (Oxphos) metabolism. Metformin, a biguanide commonly used for type 2 diabetes, inhibited the most relevant features of acid-induced phenotype, including EMT and Oxphos. When we tested effects of lactic acidosis, to verify whether sodium lactate might have additional effects on acidic melanoma cells, we found that EMT and Oxphos also characterized lactic acid-treated cells. An increased level of motility was the only gained property of lactic acidic-exposed melanoma cells. Metformin treatment inhibited both EMT markers and Oxphos and, when its concentration raised to 10 mM, it induced a striking inhibition of proliferation and colony formation of acidic melanoma cells, both grown in protons enriched medium or lactic acidosis. Thus, our study provides the first evidence that metformin may target either proton or lactic acidosis-exposed melanoma cells inhibiting EMT and Oxphox metabolism. These findings disclose a new potential rationale of metformin addition to advanced melanoma therapy, e.g. targeting acidic cell subpopulation.

  14. Micronucleus frequency in exfoliated buccal cells from hairdresser who expose to hair products

    Directory of Open Access Journals (Sweden)

    Koh Hui Yee

    2015-06-01

    Full Text Available Background: Hairdresser is one of the fastest growing occupations in today’s society. Hairdresser help styling, cutting, colouring, perming, curling, straightening hair and various treatment to customer. Somehow, hairdresser are constantly exposed to chemical substances such as aromatic amines, hydrogen peroxide, thioglycolic acid, formaldehyde in hair products which can cause damage to human’s genome. Micronucleus is one of the effective biomarker for processes associated with the induction of DNA damage. Purpose: The aim of this study was to determine the micronucleus frequencies in buccal mucosa epithelial cells of hairdresser who were exposed to chemical of hair products. Method: This study was conducted on twenty female subjects, who were divided into 2 groups: exposed and non-exposed (control group. All subjects recruited were working in the same beauty salon. Buccal cells were obtained from each individual by using cytobrush. The cells were stained with modified Feulgen-Ronssenback method and counting of micronucleus per 1000 cell was done under light microscope. The data were analyzed using independent t-test and one-way Anova (p<0.05. Result: The result showed a significant difference in micronucleus frequency between 2 groups. There were a significantly increase of micronucleus frequency in hairdressers and increase of  micronucleus frequency with the longer duration of exposure. Conclusion: It concluded that the chemical substances of hair products had affected the micronucleus frequency ofthe epithelial cells in buccal mucosa of hairdressers.

  15. The ATM and ATR inhibitors CGK733 and caffeine suppress cyclin D1 levels and inhibit cell proliferation

    Directory of Open Access Journals (Sweden)

    Sunnerhagen Per

    2009-11-01

    Full Text Available Abstract The ataxia telangiectasia mutated (ATM and the ATM- related (ATR kinases play a central role in facilitating the resistance of cancer cells to genotoxic treatment regimens. The components of the ATM and ATR regulated signaling pathways thus provide attractive pharmacological targets, since their inhibition enhances cellular sensitivity to chemo- and radiotherapy. Caffeine as well as more specific inhibitors of ATM (KU55933 or ATM and ATR (CGK733 have recently been shown to induce cell death in drug-induced senescent tumor cells. Addition of these agents to cancer cells previously rendered senescent by exposure to genotoxins suppressed the ATM mediated p21 expression required for the survival of these cells. The precise molecular pharmacology of these agents however, is not well characterized. Herein, we report that caffeine, CGK733, and to a lesser extent KU55933, inhibit the proliferation of otherwise untreated human cancer and non-transformed mouse fibroblast cell lines. Exposure of human cancer cell lines to caffeine and CGK733 was associated with a rapid decline in cyclin D1 protein levels and a reduction in the levels of both phosphorylated and total retinoblastoma protein (RB. Our studies suggest that observations based on the effects of these compounds on cell proliferation and survival must be interpreted with caution. The differential effects of caffeine/CGK733 and KU55933 on cyclin D1 protein levels suggest that these agents will exhibit dissimilar molecular pharmacological profiles.

  16. HMGA2 Inhibits Apoptosis through Interaction with ATR-CHK1 Signaling Complex in Human Cancer Cells

    Directory of Open Access Journals (Sweden)

    Suchitra Natarajan

    2013-03-01

    Full Text Available The non-histone chromatin binding protein high mobility group AT-hook 2 (HMGA2 is expressed in stem cells and many cancer cells, including tumor initiating cells, but not translated in normal human somatic cells. The presence of HMGA2 is correlated with advanced neoplastic disease and poor prognosis for patients. We had previously demonstrated a role of HMGA2 in DNA repair pathways. In the present study, we employed different human tumor cell models with endogenous and exogenous expression of HMGA2 and show that upon DNA damage, the presence of HMGA2 caused an increased and sustained phosphorylation of the ataxia telangiectasia and Rad3-related kinase (ATR and its downstream target checkpoint kinase 1 (CHK1. The presence of activated pCHK1Ser296 coincided with prolonged G2/M block and increased tumor cell survival, which was enhanced further in the presence of HMGA2. Our study, thus, identifies a novel relationship between the ATR-CHK1 DNA damage response pathway and HMGA2, which may support the DNA repair function of HMGA2 in cancer cells. Furthermore, our data provide a rationale for the use of inhibitors to ATR or CHK1 and HMGA2 in the treatment of HMGA2-positive human cancer cells.

  17. BCG vaccination induces HIV target cell activation in HIV-exposed infants in a randomized trial.

    Science.gov (United States)

    Gasper, Melanie A; Hesseling, Anneke C; Mohar, Isaac; Myer, Landon; Azenkot, Tali; Passmore, Jo-Ann S; Hanekom, Willem; Cotton, Mark F; Crispe, I Nicholas; Sodora, Donald L; Jaspan, Heather B

    2017-04-06

    BACKGROUND. Bacillus Calmette-Guérin (BCG) vaccine is administered at birth to protect infants against tuberculosis throughout Africa, where most perinatal HIV-1 transmission occurs. We examined whether BCG vaccination alters the levels of activated HIV target T cells in HIV-exposed South African infants. METHODS. HIV-exposed infants were randomized to receive routine (at birth) or delayed (at 8 weeks) BCG vaccination. Activated and CCR5-expressing peripheral blood CD4+ T cell, monocyte, and NK cell frequencies were evaluated by flow cytometry and immune gene expression via PCR using Biomark (Fluidigm). RESULTS. Of 149 infants randomized, 92% (n = 137) were retained at 6 weeks: 71 in the routine BCG arm and 66 in the delayed arm. Routine BCG vaccination led to a 3-fold increase in systemic activation of HIV target CD4+CCR5+ T cells (HLA-DR+CD38+) at 6 weeks (0.25% at birth versus 0.08% in delayed vaccination groups; P = 0.029), which persisted until 8 weeks of age when the delayed arm was vaccinated. Vaccination of the infants in the delayed arm at 8 weeks resulted in a similar increase in activated CD4+CCR5+ T cells. The increase in activated T cells was associated with increased levels of MHC class II transactivator (CIITA), IL12RB1, and IFN-α1 transcripts within peripheral blood mononuclear cells but minimal changes in innate cells. CONCLUSION. BCG vaccination induces immune changes in HIV-exposed infants, including an increase in the proportion of activated CCR5+CD4+ HIV target cells. These findings provide insight into optimal BCG vaccine timing to minimize the risks of HIV transmissions to exposed infants while preserving potential benefits conferred by BCG vaccination. TRIAL REGISTRATION. ClinicalTrials.gov NCT02062580. FUNDING. This trial was sponsored by the Elizabeth Glaser Pediatric AIDS Foundation (MV-00-9-900-01871-0-00) and the Thrasher Foundation (NR-0095); for details, see Acknowledgments.

  18. Superoxide dismutase activity and chromosome damage in cultured chromosome instability syndrome cells.

    Science.gov (United States)

    Lee, K H; Abe, S; Yanabe, Y; Matsuda, I; Yoshida, M C

    1990-07-01

    The basal levels of superoxide dismutase (SOD) activity and chromosome aberration (CA) and sister-chromatid exchange (SCE) frequencies were examined in cultured fibroblasts or Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCLs). These cells were derived from patients with chromosome instability syndromes (CISs) including Bloom's syndrome (BS), Fanconi's anemia (FA) and ataxia telangiectasia (AT). Embryonal fibroblasts and LCLs from normal subjects served as controls. Although LCLs tended to exhibit a higher SOD level than fibroblasts due to an elevation of Cu/Zn-SOD activity, BS and FA fibroblasts with increased frequencies of CAs and/or SCEs showed abnormally elevated SOD activity due to the manifold increase of Mn-SOD levels compared with control cells. However, BS and AT LCLs with almost control levels of CA and SCE frequencies showed no, or a slightly elevated, SOD activity, suggesting a possible selection of such cells during EBV transformation. The observed parallelism between the SOD activity and the cytogenetic manifestation may imply an involvement of active oxygen species, especially superoxide radicals, in the increased chromosome damage of CIS cells.

  19. NPRL-Z-1, as a new topoisomerase II poison, induces cell apoptosis and ROS generation in human renal carcinoma cells.

    Science.gov (United States)

    Wu, Szu-Ying; Pan, Shiow-Lin; Xiao, Zhi-Yan; Hsu, Jui-Ling; Chen, Mei-Chuan; Lee, Kuo-Hsiung; Teng, Che-Ming

    2014-01-01

    NPRL-Z-1 is a 4β-[(4"-benzamido)-amino]-4'-O-demethyl-epipodophyllotoxin derivative. Previous reports have shown that NPRL-Z-1 possesses anticancer activity. Here NPRL-Z-1 displayed cytotoxic effects against four human cancer cell lines (HCT 116, A549, ACHN, and A498) and exhibited potent activity in A498 human renal carcinoma cells, with an IC50 value of 2.38 µM via the MTT assay. We also found that NPRL-Z-1 induced cell cycle arrest in G1-phase and detected DNA double-strand breaks in A498 cells. NPRL-Z-1 induced ataxia telangiectasia-mutated (ATM) protein kinase phosphorylation at serine 1981, leading to the activation of DNA damage signaling pathways, including Chk2, histone H2AX, and p53/p21. By ICE assay, the data suggested that NPRL-Z-1 acted on and stabilized the topoisomerase II (TOP2)-DNA complex, leading to TOP2cc formation. NPRL-Z-1-induced DNA damage signaling and apoptotic death was also reversed by TOP2α or TOP2β knockdown. In addition, NPRL-Z-1 inhibited the Akt signaling pathway and induced reactive oxygen species (ROS) generation. These results demonstrated that NPRL-Z-1 appeared to be a novel TOP2 poison and ROS generator. Thus, NPRL-Z-1 may present a significant potential anticancer candidate against renal carcinoma.

  20. AZD6738, A Novel Oral Inhibitor of ATR, Induces Synthetic Lethality with ATM Deficiency in Gastric Cancer Cells.

    Science.gov (United States)

    Min, Ahrum; Im, Seock-Ah; Jang, Hyemin; Kim, Seongyeong; Lee, Miso; Kim, Debora Keunyoung; Yang, Yaewon; Kim, Hee-Jun; Lee, Kyung-Hun; Kim, Jin Won; Kim, Tae-Yong; Oh, Do-Youn; Brown, Jeff; Lau, Alan; O'Connor, Mark J; Bang, Yung-Jue

    2017-04-01

    Ataxia telangiectasia and Rad3-related (ATR) can be considered an attractive target for cancer treatment due to its deleterious effect on cancer cells harboring a homologous recombination defect. The aim of this study was to investigate the potential use of the ATR inhibitor, AZD6738, to treat gastric cancer.In SNU-601 cells with dysfunctional ATM, AZD6738 treatment led to an accumulation of DNA damage due to dysfunctional RAD51 foci formation, S phase arrest, and caspase 3-dependent apoptosis. In contrast, SNU-484 cells with functional ATM were not sensitive to AZD6738. Inhibition of ATM in SNU-484 cells enhanced AZD6738 sensitivity to a level comparable with that observed in SNU-601 cells, showing that activation of the ATM-Chk2 signaling pathway attenuates AZD6738 sensitivity. In addition, decreased HDAC1 expression was found to be associated with ATM inactivation in SNU-601 cells, demonstrating the interaction between HDAC1 and ATM can affect sensitivity to AZD6738. Furthermore, in an in vivo tumor xenograft mouse model, AZD6738 significantly suppressed tumor growth and increased apoptosis.These findings suggest synthetic lethality between ATR inhibition and ATM deficiency in gastric cancer cells. Further clinical studies on the interaction between AZD 6738 and ATM deficiency are warranted to develop novel treatment strategies for gastric cancer. Mol Cancer Ther; 16(4); 566-77. ©2017 AACR. ©2017 American Association for Cancer Research.

  1. Ultrastructure of cells of Ulmus americana cultured in vitro and exposed to the culture filtrate of Ceratocystis ulmi

    Science.gov (United States)

    Paula M. Pijut; R. Daniel Lineberger; Subhash C. Domir; Jann M. Ichida; Charles R. Krause

    1990-01-01

    Calli of American elm susceptible and resistant to Dutch elm disease were exposed to a culture filtrate of a pathogenic isolate of Ceratocystis ulmi. Cells from untreated tissue exhibited typical internal composition associated with healthy, actively growing cells. All cells exposed to culture filtrate showed appreciable ultrastructural changes....

  2. Targeting Werner syndrome protein sensitizes U-2 OS osteosarcoma cells to selenium-induced DNA damage response and necrotic death

    DEFF Research Database (Denmark)

    Cheng, Wen-Hsing; Wu, Ryan T Y; Wu, Min

    2012-01-01

    , but promoted recovery from the MSeA-induced DNA damage. Taken together, WRN protects U-2 OS osteosarcoma cells against MSeA-induced cytotoxicity, suggesting that oxidative DNA repair pathway is a promising target for improving the efficacy of selenium on tumor suppression....... in mouse models of cancer. To test the hypothesis that targeting WRN can potentiate selenium toxicity in cancer cells, isogenic WRN small hairpin RNA (shRNA) and control shRNA U-2 OS osteosarcoma cells were treated with MSeA for 2d, followed by recovery for up to 7d. WRN deficiency sensitized U-2 OS cells...... to MSeA-induced necrotic death. Co-treatment with the ataxia-telangiectasia mutated (ATM) kinase inhibitor KU55933 desensitized the control shRNA cells, but not WRN shRNA cells, to MSeA treatment. WRN did not affect MSeA-induced ATM phosphorylation on Ser-1981 or H2A.X phosphorylation on Ser-139...

  3. Activation of Holliday junction recognizing protein involved in the chromosomal stability and immortality of cancer cells.

    Science.gov (United States)

    Kato, Tatsuya; Sato, Nagato; Hayama, Satoshi; Yamabuki, Takumi; Ito, Tomoo; Miyamoto, Masaki; Kondo, Satoshi; Nakamura, Yusuke; Daigo, Yataro

    2007-09-15

    We identified a novel gene HJURP (Holliday junction-recognizing protein) whose activation seemed to play a pivotal role in the immortality of cancer cells. HJURP was considered a possible downstream target for ataxia telangiectasia mutated signaling, and its expression was increased by DNA double-strand breaks (DSB). HJURP was involved in the homologous recombination pathway in the DSB repair process through interaction with hMSH5 and NBS1, which is a part of the MRN protein complex. HJURP formed nuclear foci in cells at S phase and those subjected to DNA damage. In vitro assays implied that HJURP bound directly to the Holliday junction and rDNA arrays. Treatment of cancer cells with small interfering RNA (siRNA) against HJURP caused abnormal chromosomal fusions and led to genomic instability and senescence. In addition, HJURP overexpression was observed in a majority of lung cancers and was associated with poor prognosis as well. We suggest that HJURP is an indispensable factor for chromosomal stability in immortalized cancer cells and is a potential novel therapeutic target for the development of anticancer drugs.

  4. In vitro metabolism study of normal and tumor cells when exposed to red LED light

    Science.gov (United States)

    Stolbovskaya, Olga V.; Khairullin, Radik M.; Saenko, Yuri V.; Krasnikova, Ekaterina S.; Krasnikov, Aleksandr V.; Fomin, Aleksandr A.; Skaptsov, Aleksandr A.

    2016-04-01

    This work presents the results of studying the mitochondrial membrane potential, intracellular ROS, peculiarities of the cell cycle of cancer cells HCT-116 and the normal line of CHO cells when exposed to the red LED light with a wavelength range of 0.620-0.680 μm. A dose-dependent increase in mitochondrial membrane potential and intracellular ROS concentration in cancer cells HCT-116 was established. In normal CHO cell line a dose-dependent reduction of mitochondrial membrane potential and dose-dependent increase in intracellular ROS occur. It has been shown that the sensitivity of the studied cell lines to the red light depends on the stage of the cell cycle.

  5. Host cell reactivation of uv- and X-ray-damaged herpes simplex virus by Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Henderson, E.E. (Temple Univ. School of Medicine, Philadelphia, PA); Long, W.K.

    1981-12-01

    The efficacy of using an infected centers assay, employing herpes simplex virus-infected, Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) as components, to study host cell reactivation has been explored. Herpes simplex virus type 1 (HSV-1) was shown through the infected centers assay to have detectable but varying ability to lytically infect LCLs established from chromosomal breakage syndromes or closely related genetic disorders. The rate of HSV inactivation by ultraviolet (uv) irradiation was faster in LCLs established from Cockaynes's syndrome than in normal LCLs, and faster still in LCLs established from xeroderma pigmentosum. These results indicate that Cockayne's syndrome, while having what appears to be quantitatively normal levels of uv-induced DNA repair replication, shows decreased ability to host cell reactivated uv-damaged HSV. In direct contrast, X-irradiated HSV showed identical survival when assayed on normal LCLs or LCLs established from ataxia telangiectasia showing increased sensitivity to X irradiation as measured by colony formation. Through the infected centers assay, it has also been possible to demonstrate low levels of multiplicity reactivation of mutagen-damaged HSV in permanently proliferating LCLs.

  6. Cellular and exosome mediated molecular defense mechanism in bovine granulosa cells exposed to oxidative stress.

    Science.gov (United States)

    Saeed-Zidane, Mohammed; Linden, Lea; Salilew-Wondim, Dessie; Held, Eva; Neuhoff, Christiane; Tholen, Ernst; Hoelker, Michael; Schellander, Karl; Tesfaye, Dawit

    2017-01-01

    Various environmental insults including diseases, heat and oxidative stress could lead to abnormal growth, functions and apoptosis in granulosa cells during ovarian follicle growth and oocyte maturation. Despite the fact that cells exposed to oxidative stress are responding transcriptionally, the potential release of transcripts associated with oxidative stress response into extracellular space through exosomes is not yet determined. Therefore, here we aimed to investigate the effect of oxidative stress in bovine granulosa cells in vitro on the cellular and exosome mediated defense mechanisms. Bovine granulosa cells were aspirated from ovarian follicles and cultured in DMEM/F-12 Ham culture medium supplemented with 10% exosome-depleted fetal bovine serum. In the first experiment sub-confluent cells were treated with 5 μM H2O2 for 40 min to induce oxidative stress. Thereafter, cells were subjected to ROS and mitochondrial staining, cell proliferation and cell cycle assays. Furthermore, gene and protein expression analysis were performed in H2O2-challenged versus control group 24 hr post-treatment using qRT-PCR and immune blotting or immunocytochemistry assay, respectively. Moreover, exosomes were isolated from spent media using ultracentrifugation procedure, and subsequently used for RNA isolation and qRT-PCR. In the second experiment, exosomes released by granulosa cells under oxidative stress (StressExo) or those released by granulosa cells without oxidative stress (NormalExo) were co-incubated with bovine granulosa cells in vitro to proof the potential horizontal transfer of defense molecules from exosomes to granulosa cells and investigate any phenotype changes. Exposure of bovine granulosa cells to H2O2 induced the accumulation of ROS, reduced mitochondrial activity, increased expression of Nrf2 and its downstream antioxidant genes (both mRNA and protein), altered the cell cycle transitions and induced cellular apoptosis. Granulosa cells exposed to oxidative

  7. ATM kinase sustains breast cancer stem-like cells by promoting ATG4C expression and autophagy.

    Science.gov (United States)

    Antonelli, Martina; Strappazzon, Flavie; Arisi, Ivan; Brandi, Rossella; D'Onofrio, Mara; Sambucci, Manolo; Manic, Gwenola; Vitale, Ilio; Barilà, Daniela; Stagni, Venturina

    2017-03-28

    The efficacy of Ataxia-Telangiectasia Mutated (ATM) kinase signalling inhibition in cancer therapy is tempered by the identification of new emerging functions of ATM, which suggests that the role of this protein in cancer progression is complex. We recently demonstrated that this tumor suppressor gene could act as tumor promoting factor in HER2 (Human Epidermal Growth Factor Receptor 2) positive breast cancer. Herein we put in evidence that ATM expression sustains the proportion of cells with a stem-like phenotype, measured as the capability to form mammospheres, independently of HER2 expression levels. Transcriptomic analyses revealed that, in mammospheres, ATM modulates the expression of cell cycle-, DNA repair- and autophagy-related genes. Among these, the silencing of the autophagic gene, autophagy related 4C cysteine peptidase (ATG4C), impairs mammosphere formation similarly to ATM depletion. Conversely, ATG4C ectopic expression in cells silenced for ATM expression, rescues mammospheres growth. Finally, tumor array analyses, performed using public data, identify a significant correlation between ATM and ATG4C expression levels in all human breast cancer subtypes, except for the basal-like one.Overall, we uncover a new connection between ATM kinase and autophagy regulation in breast cancer. We demonstrate that, in breast cancer cells, ATM and ATG4C are essential drivers of mammosphere formation, suggesting that their targeting may improve current approaches to eradicate breast cancer cells with a stem-like phenotype.

  8. Evaluation of ATM heterozygous mutations underlying individual differences in radiosensitivity using genome editing in human cultured cells.

    Science.gov (United States)

    Royba, Ekaterina; Miyamoto, Tatsuo; Natsuko Akutsu, Silvia; Hosoba, Kosuke; Tauchi, Hiroshi; Kudo, Yoshiki; Tashiro, Satoshi; Yamamoto, Takashi; Matsuura, Shinya

    2017-07-20

    Ionizing radiation (IR) induces DNA double-strand breaks (DSBs), which are an initial step towards chromosomal aberrations and cell death. It has been suggested that there are individual differences in radiosensitivity within human populations, and that the variations in DNA repair genes might determine this heterogeneity. However, it is difficult to quantify the effect of genetic variants on the individual differences in radiosensitivity, since confounding factors such as smoking and the diverse genetic backgrounds within human populations affect radiosensitivity. To precisely quantify the effect of a genetic variation on radiosensitivity, we here used the CRISPR-ObLiGaRe (Obligate Ligation-Gated Recombination) method combined with the CRISPR/Cas9 system and a nonhomologous end joining (NHEJ)-mediated knock-in technique in human cultured cells with a uniform genetic background. We generated ATM heterozygous knock-out (ATM +/- ) cell clones as a carrier model of a radiation-hypersensitive autosomal-recessive disorder, ataxia-telangiectasia (A-T). Cytokinesis-blocked micronucleus assay and chromosome aberration assay showed that the radiosensitivity of ATM +/- cell clones was significantly higher than that of ATM +/+ cells, suggesting that ATM gene variants are indeed involved in determining individual radiosensitivity. Importantly, the differences in radiosensitivity among the same genotype clones were small, unlike the individual differences in fibroblasts derived from A-T-affected family members.

  9. The transcriptome of HIV-1 infected intestinal CD4+ T cells exposed to enteric bacteria.

    Science.gov (United States)

    Yoder, Alyson C; Guo, Kejun; Dillon, Stephanie M; Phang, Tzu; Lee, Eric J; Harper, Michael S; Helm, Karen; Kappes, John C; Ochsenbauer, Christina; McCarter, Martin D; Wilson, Cara C; Santiago, Mario L

    2017-02-01

    Global transcriptome studies can help pinpoint key cellular pathways exploited by viruses to replicate and cause pathogenesis. Previous data showed that laboratory-adapted HIV-1 triggers significant gene expression changes in CD4+ T cell lines and mitogen-activated CD4+ T cells from peripheral blood. However, HIV-1 primarily targets mucosal compartments during acute infection in vivo. Moreover, early HIV-1 infection causes extensive depletion of CD4+ T cells in the gastrointestinal tract that herald persistent inflammation due to the translocation of enteric microbes to the systemic circulation. Here, we profiled the transcriptome of primary intestinal CD4+ T cells infected ex vivo with transmitted/founder (TF) HIV-1. Infections were performed in the presence or absence of Prevotella stercorea, a gut microbe enriched in the mucosa of HIV-1-infected individuals that enhanced both TF HIV-1 replication and CD4+ T cell death ex vivo. In the absence of bacteria, HIV-1 triggered a cellular shutdown response involving the downregulation of HIV-1 reactome genes, while perturbing genes linked to OX40, PPAR and FOXO3 signaling. However, in the presence of bacteria, HIV-1 did not perturb these gene sets or pathways. Instead, HIV-1 enhanced granzyme expression and Th17 cell function, inhibited G1/S cell cycle checkpoint genes and triggered downstream cell death pathways in microbe-exposed gut CD4+ T cells. To gain insights on these differential effects, we profiled the gene expression landscape of HIV-1-uninfected gut CD4+ T cells exposed to bacteria. Microbial exposure upregulated genes involved in cellular proliferation, MAPK activation, Th17 cell differentiation and type I interferon signaling. Our findings reveal that microbial exposure influenced how HIV-1 altered the gut CD4+ T cell transcriptome, with potential consequences for HIV-1 susceptibility, cell survival and inflammation. The HIV-1- and microbe-altered pathways unraveled here may serve as a molecular blueprint

  10. The transcriptome of HIV-1 infected intestinal CD4+ T cells exposed to enteric bacteria

    Science.gov (United States)

    Dillon, Stephanie M.; Phang, Tzu; Lee, Eric J.; Helm, Karen; Kappes, John C.; McCarter, Martin D.

    2017-01-01

    Global transcriptome studies can help pinpoint key cellular pathways exploited by viruses to replicate and cause pathogenesis. Previous data showed that laboratory-adapted HIV-1 triggers significant gene expression changes in CD4+ T cell lines and mitogen-activated CD4+ T cells from peripheral blood. However, HIV-1 primarily targets mucosal compartments during acute infection in vivo. Moreover, early HIV-1 infection causes extensive depletion of CD4+ T cells in the gastrointestinal tract that herald persistent inflammation due to the translocation of enteric microbes to the systemic circulation. Here, we profiled the transcriptome of primary intestinal CD4+ T cells infected ex vivo with transmitted/founder (TF) HIV-1. Infections were performed in the presence or absence of Prevotella stercorea, a gut microbe enriched in the mucosa of HIV-1-infected individuals that enhanced both TF HIV-1 replication and CD4+ T cell death ex vivo. In the absence of bacteria, HIV-1 triggered a cellular shutdown response involving the downregulation of HIV-1 reactome genes, while perturbing genes linked to OX40, PPAR and FOXO3 signaling. However, in the presence of bacteria, HIV-1 did not perturb these gene sets or pathways. Instead, HIV-1 enhanced granzyme expression and Th17 cell function, inhibited G1/S cell cycle checkpoint genes and triggered downstream cell death pathways in microbe-exposed gut CD4+ T cells. To gain insights on these differential effects, we profiled the gene expression landscape of HIV-1-uninfected gut CD4+ T cells exposed to bacteria. Microbial exposure upregulated genes involved in cellular proliferation, MAPK activation, Th17 cell differentiation and type I interferon signaling. Our findings reveal that microbial exposure influenced how HIV-1 altered the gut CD4+ T cell transcriptome, with potential consequences for HIV-1 susceptibility, cell survival and inflammation. The HIV-1- and microbe-altered pathways unraveled here may serve as a molecular blueprint

  11. Role of Mre11 in chromosomal nonhomologous end joining in mammalian cells.

    Science.gov (United States)

    Rass, Emilie; Grabarz, Anastazja; Plo, Isabelle; Gautier, Jean; Bertrand, Pascale; Lopez, Bernard S

    2009-08-01

    Here we have used an intrachromosomal substrate to monitor the end joining of distant ends, which leads to DNA rearrangements in mammalian cells. We show that silencing Mre11 reduces the efficiency of nonhomologous end joining (NHEJ), affecting both the canonical and alternative pathways, partly in a manner that is independent of the ataxia-telangiectasia mutated kinase (ATM). Silencing of Rad50 or CtIP decreases end-joining efficiency in the same pathway as Mre11. In cells defective for Xrcc4, the MRE11-RAD50-NBS1 (MRN) complex inhibitor MIRIN decreases end-joining frequencies, demonstrating a role for MRN in alternative NHEJ. Consistently, MIRIN sensitizes both complemented and NHEJ-defective cells to ionizing radiation. Conversely, overexpression of Mre11 stimulates the resection of single-stranded DNA and increases alternative end joining, through a mechanism that requires Mre11's nuclease activity, but in an ATM-independent manner. These data demonstrate that, in addition to its role in ATM activation, Mre11 can favor alternative NHEJ through its nuclease activity.

  12. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke.

    Science.gov (United States)

    Berman, Reena; Jiang, Di; Wu, Qun; Chu, Hong Wei

    2016-01-01

    Human rhinovirus (HRV) infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS) increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT) reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air-liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS) with or without HRV-16 (5×10(4) 50% Tissue Culture Infective Dose [TCID50]/transwell) infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection.

  13. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke

    Directory of Open Access Journals (Sweden)

    Berman R

    2016-06-01

    Full Text Available Reena Berman, Di Jiang, Qun Wu, Hong Wei Chu Department of Medicine, National Jewish Health, Denver, CO, USA Abstract: Human rhinovirus (HRV infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air–liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS with or without HRV-16 (5×104 50% Tissue Culture Infective Dose [TCID50]/transwell infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection. Keywords: α1-antitrypsin, rhinovirus, COPD, cigarette smoke, ICAM-1

  14. Identification of signaling pathways mediating cell cycle arrest and apoptosis induced by Porphyromonas gingivalis in human trophoblasts.

    Science.gov (United States)

    Inaba, Hiroaki; Kuboniwa, Masae; Sugita, Hideyuki; Lamont, Richard J; Amano, Atsuo

    2012-08-01

    Epidemiological and interventional studies of humans have revealed a close association between periodontal diseases and preterm delivery of low-birth-weight infants. Porphyromonas gingivalis, a periodontal pathogen, can translocate to gestational tissues following oral-hematogenous spread. We previously reported that P. gingivalis invades extravillous trophoblast cells (HTR-8) derived from the human placenta and inhibits proliferation through induction of arrest in the G(1) phase of the cell cycle. The purpose of the present study was to identify signaling pathways mediating cellular impairment caused by P. gingivalis. Following P. gingivalis infection, the expression of Fas was induced and p53 accumulated, responses consistent with response to DNA damage. Ataxia telangiectasia- and Rad3-related kinase (ATR), an essential regulator of DNA damage checkpoints, was shown to be activated together with its downstream signaling molecule Chk2, while the p53 degradation-related protein MDM2 was not induced. The inhibition of ATR prevented both G(1) arrest and apoptosis caused by P. gingivalis in HTR-8 cells. In addition, small interfering RNA (siRNA) knockdown of p53 abrogated both G(1) arrest and apoptosis. The regulation of apoptosis was associated with Ets1 activation. HTR-8 cells infected with P. gingivalis exhibited activation of Ets1, and knockdown of Ets1 with siRNA diminished both G(1) arrest and apoptosis. These results suggest that P. gingivalis activates cellular DNA damage signaling pathways that lead to G(1) arrest and apoptosis in trophoblasts.

  15. Static electric fields interfere in the viability of cells exposed to ionising radiation.

    Science.gov (United States)

    Arruda-Neto, João D T; Friedberg, Errol C; Bittencourt-Oliveira, Maria C; Cavalcante-Silva, Erika; Schenberg, Ana C G; Rodrigues, Tulio E; Garcia, Fermin; Louvison, Monica; Paula, Claudete R; Mesa, Joel; Moron, Michelle M; Maria, Durvanei A; Genofre, Godofredo C

    2009-04-01

    The interference of electric fields (EF) with biological processes is an issue of considerable interest. No studies have as yet been reported on the combined effect of EF plus ionising radiation. Here we report studies on this combined effect using the prokaryote Microcystis panniformis, the eukaryote Candida albicans and human cells. Cultures of Microcystis panniformis (Cyanobacteria) in glass tubes were irradiated with doses in the interval 0.5-5 kGy, using a (60)Co gamma source facility. Samples irradiated with 3 kGy were exposed for 2 h to a 20 V . cm(-1) static electric field and viable cells were enumerated. Cultures of Candida albicans were incubated at 36 degrees C for 20 h, gamma-irradiated with doses from 1-4 kGy, and submitted to an electric field of 180 V . cm(-1). Samples were examined under a fluorescence microscope and the number of unviable (red) and viable (apple green fluorescence) cells was determined. For crossing-check purposes, MRC5 strain of lung cells were irradiated with 2 Gy, exposed to an electric field of 1250 V/cm, incubated overnight with the anti-body anti-phospho-histone H2AX and examined under a fluorescence microscope to quantify nuclei with gamma-H2AX foci. In cells exposed to EF, death increased substantially compared to irradiation alone. In C. albicans we observed suppression of the DNA repair shoulder. The effect of EF in growth of M. panniformis was substantial; the number of surviving cells on day-2 after irradiation was 12 times greater than when an EF was applied. By the action of a static electric field on the irradiated MRC5 cells the number of nuclei with gamma-H2AX foci increased 40%, approximately. Application of an EF following irradiation greatly increases cell death. The observation that the DNA repair shoulder in the survival curve of C. albicans is suppressed when cells are exposed to irradiation + EF suggests that EF likely inactivate cellular recovering processes. The result for the number of nuclei with gamma

  16. Apoptosis and necroptosis are induced in rainbow trout cell lines exposed to cadmium

    Energy Technology Data Exchange (ETDEWEB)

    Krumschnabel, Gerhard, E-mail: Gerhard.Krumschnabel@i-med.ac.at [Division of Developmental Immunology, Biocenter, Medical University Innsbruck, Fritz-Preglstr. 3, Innsbruck (Austria); Ebner, Hannes L.; Hess, Michael W. [Division of Histology and Embryology, Medical University Innsbruck, Innsbruck (Austria); Villunger, Andreas [Division of Developmental Immunology, Biocenter, Medical University Innsbruck, Fritz-Preglstr. 3, Innsbruck (Austria)

    2010-08-01

    Cadmium is an important environmental toxicant that can kill cells. A number of studies have implicated apoptosis as well as necrosis and, most recently, a form of programmed necrosis termed necroptosis in the process of cadmium-mediated toxicity, but the exact mechanism remains ill-defined and may depend on the affected cell type. This study investigated which mode of cell death may be responsible for cell death induction in cadmium-exposed trout cell lines from gill and liver and if this cell death was sensitive to inhibitors of necroptosis or apoptosis, respectively. It was observed that intermediate levels of cadmium that killed approximately 50% of the cells over 96-120 h of exposure caused cell death that morphologically resembled apoptosis and was associated with an increase of apoptotic markers such as the number of cells with diminished DNA content (sub-G1 cells), condensed or fragmented nuclei, and elevation of caspase-3 activity. At the same time, however, cells also lost plasma membrane integrity, as indicated by uptake of propidium iodide, showed a decrease of ATP levels and mitochondrial membrane potential, and displayed cell swelling, signs associated with secondary necrosis, or equally possible, necroptotic cell death. Importantly, many of these alterations were at least partly inhibited by the necroptosis inhibitor necrostatin-1 and were to a lesser extent also sensitive to the pan-caspase inhibitor zVAD-fmk, indicating that multiple modes of cell death are concurrently induced in cadmium-exposed trout cells, including necroptosis and apoptosis. Cell death appeared to lack concurrent radical formation, consistent with genetically regulated necroptotic cell death, but was characterized by the rapid induction of DNA damage markers, and the early onset of disintegration of the Golgi complex. Comparative experiments evaluating copper-toxicity indicated that in comparison to cadmium much higher concentrations of this metal were required to induce cell

  17. Functional activation of T cells by dendritic cells and macrophages exposed to the intracellular parasite Neospora caninum.

    Science.gov (United States)

    Dion, Sarah; Germon, Stéphanie; Guiton, Rachel; Ducournau, Céline; Dimier-Poisson, Isabelle

    2011-05-01

    Neospora caninum is an intracellular protozoan pathogen that causes abortion in cattle. We studied how the interaction between murine conventional dendritic cells or macrophages and N. caninum influences the generation of cell-mediated immunity against the parasite. We first explored the invasion and survival ability of N. caninum in dendritic cells and macrophages. We observed that protozoa rapidly invaded and proliferated into these two cell populations. We then investigated how Neospora-exposed macrophages or dendritic cells distinguish between viable and non-viable (heat-killed tachyzoites and antigenic extract) parasites. Viable tachyzoites and antigenic extract, but not killed parasites, altered the phenotype of immature dendritic cells. Dendritic cells infected with viable parasites down-regulated the expression of MHC-II, CD40, CD80 and CD86 whereas dendritic cells exposed to N. caninum antigenic extract up-regulated the expression of MHC-II and CD40 and down-regulated CD80 and CD86 expression. Moreover, only viable tachyzoites and antigenic extract induced IL-12 synthesis by dendritic cells. MHC-II expression was up-regulated and CD86 expression was down-regulated at the surface of macrophages, regardless of the parasitic form was encountered. However, IL-12 secretion by macrophages was only observed under conditions using viable and heat-killed parasite. We then analysed how macrophages and dendritic cells were involved in inducing T-cell responses. T lymphocyte IFN-γ-secretion in correlation with IL-12 production occurred after interactions between T cells and dendritic cells exposed to viable tachyzoites or antigenic extract. By contrast, for macrophages IFN-γ production was IL-12-independent and only occurred after interactions between T cells and macrophages exposed to antigenic extract. Thus, N. caninum-induced activation of murine dendritic cells, but not that of macrophages, was associated with T cell IFN-γ production after IL-12 secretion

  18. Frequency patterns of T-cell exposed motifs in immunoglobulin heavy chain peptides presented by MHCs

    Directory of Open Access Journals (Sweden)

    Robert D. Bremel

    2014-10-01

    Full Text Available Immunoglobulins are highly diverse protein sequences that are processed and presented to T-cells by B-cells and other antigen presenting cells. We examined a large dataset of immunoglobulin heavy chain variable regions (IGHV to assess the diversity of T-cell exposed motifs (TCEM. TCEM comprise those amino acids in a MHC-bound peptide which face outwards, surrounded by the MHC histotope, and which engage the T-cell receptor. Within IGHV there is a distinct pattern of predicted MHC class II binding and a very high frequency of re-use of the TCEMs. The re-use frequency indicates that only a limited number of different cognate T-cells are required to engage many different clonal B-cells. The amino acids in each outward-facing TCEM are intercalated with the amino acids of inward-facing MHC groove-exposed motifs (GEM. Different GEM may have differing, allele-specific, MHC binding affinities. The intercalation of TCEM and GEM in a peptide allows for a vast combinatorial repertoire of epitopes, each eliciting a different response. Outcome of T-cell receptor binding is determined by overall signal strength, which is a function of the number of responding T-cells and the duration of engagement. Hence, the frequency of T-cell exposed motif re-use appears to be an important determinant of whether a T-cell response is stimulatory or suppressive. The frequency distribution of TCEMs implies that somatic hypermutation is followed by clonal expansion that develop along repeated pathways. The observations of TCEM and GEM derived from immunoglobulins suggest a relatively simple, yet powerful, mechanism to correlate T-cell polyspecificity, through re-use of TCEMs, with a very high degree of specificity achieved by combination with a diversity of GEMs. The frequency profile of TCEMs also points to an economical mechanism for maintaining T-cell memory, recall, and self-discrimination based on an endogenously generated profile of motifs.

  19. Mechanisms of mutagenesis in human cells exposed to 55 MeV protons

    Science.gov (United States)

    Gauny, S.; Wiese, C.; Kronenberg, A.

    2001-01-01

    Protons represent the major type of charged particle radiation in spaceflight environments. The purpose of this study was to assess mutations arising in human lymphoid cells exposed to protons. Mutations were quantitated at the thymidine kinase (TK1) locus in cell lines derived from the same donor: TK6 cells (wt TP53) and WTK1 cells (mutant TP53). WTK1 cells were much more susceptible to mutagenesis following proton exposure than TK6 cells. Intragenic deletions were observed among early-arising TK1 mutants in TK6 cells, but not in WTK1 cells where all of the mutants arose by LOH. Deletion was the predominant mode of LOH in TK6 cells, while allelic recombination was the major mode of LOH in WTK1 cells. Deletions were of variable lengths, from recombination often extended to the telomere. In summary, proton exposures elicited many types of mutations at an autosomal locus in human cells. Most involved large scale loss of genetic information, either through deletion or by recombination.

  20. Assembly and Reorientation of Stress Fibers Drives Morphological Changes to Endothelial Cells Exposed to Shear Stress

    Science.gov (United States)

    Noria, Sabrena; Xu, Feng; McCue, Shannon; Jones, Mara; Gotlieb, Avrum I.; Langille, B. Lowell

    2004-01-01

    Fluid shear stress greatly influences the biology of vascular endothelial cells and the pathogenesis of atherosclerosis. Endothelial cells undergo profound shape change and reorientation in response to physiological levels of fluid shear stress. These morphological changes influence cell function; however, the processes that produce them are poorly understood. We have examined how actin assembly is related to shear-induced endothelial cell shape change. To do so, we imposed physiological levels of shear stress on cultured endothelium for up to 96 hours and then permeabilized the cells and exposed them briefly to fluorescently labeled monomeric actin at various time points to assess actin assembly. Alternatively, monomeric actin was microinjected into cells to allow continuous monitoring of actin distribution. Actin assembly occurred primarily at the ends of stress fibers, which simultaneously reoriented to the shear axis, frequently fused with neighboring stress fibers, and ultimately drove the poles of the cells in the upstream and/or downstream directions. Actin polymerization occurred where stress fibers inserted into focal adhesion complexes, but usually only at one end of the stress fiber. Neither the upstream nor downstream focal adhesion complex was preferred. Changes in actin organization were accompanied by translocation and remodeling of cell-substrate adhesion complexes and transient formation of punctate cell-cell adherens junctions. These findings indicate that stress fiber assembly and realignment provide a novel mode by which cell morphology is altered by mechanical signals. PMID:15039210

  1. Host cell reactivation by fibroblasts from patients with pigmentary degeneration of the retina

    Energy Technology Data Exchange (ETDEWEB)

    Lytle, C.D. (Food and Drug Administration, Rockville, MD (USA)); Tarone, R.E.; Barrett, S.F.; Robbins, J.H. (National Cancer Inst., Bethesda, MD (USA)); Wirtschafter, J.D. (Minnesota Univ., Minneapolis (USA). Hospitals); Dupuy, J.M. (Quebec Univ., Laval-des-Rapides (Canada). Inst. Armand-Frappier)

    1983-05-01

    Cockayne syndrome (CS) is an autosomal recessive disease characterized by numerous clinical abnormalities including acute sun sensitivity and primary pigmentary degeneration of the retina. Cultured fibroblasts from CS patients are hypersensitive to ultraviolet radiation. Host cell reactivation of irradiated virus was studied in CS and in other diseases with retinal degeneration to evaluate repair. The survival of UV-irradiated Herpes simplex virus type 1 was determined in fibroblast lines from four normal donors, two patients with CS, one with both xeroderma pigmentosum (XP) and CS, and from several other patients with (Usher syndrome, olivopontocerebellar atrophy, retinitis pigmentosa) and without (XP, ataxia telangiectasia) primary pigmentary degeneration of the retina. The viral survival curves in all cell lines showed two components: a very sensitive initial component followed by an exponential, less sensitive component. The exponential component had greater sensitivity than normal in the case of the CS patients, the patient with both XP and CS, and the XP patient. It was proposed that patients with CS have defective repair of DNA which may be the cause of their retinal degeneration.

  2. ATM facilitates mouse gammaherpesvirus reactivation from myeloid cells during chronic infection.

    Science.gov (United States)

    Kulinski, Joseph M; Darrah, Eric J; Broniowska, Katarzyna A; Mboko, Wadzanai P; Mounce, Bryan C; Malherbe, Laurent P; Corbett, John A; Gauld, Stephen B; Tarakanova, Vera L

    2015-09-01

    Gammaherpesviruses are cancer-associated pathogens that establish life-long infection in most adults. Insufficiency of Ataxia-Telangiectasia mutated (ATM) kinase leads to a poor control of chronic gammaherpesvirus infection via an unknown mechanism that likely involves a suboptimal antiviral response. In contrast to the phenotype in the intact host, ATM facilitates gammaherpesvirus reactivation and replication in vitro. We hypothesized that ATM mediates both pro- and antiviral activities to regulate chronic gammaherpesvirus infection in an immunocompetent host. To test the proposed proviral activity of ATM in vivo, we generated mice with ATM deficiency limited to myeloid cells. Myeloid-specific ATM deficiency attenuated gammaherpesvirus infection during the establishment of viral latency. The results of our study uncover a proviral role of ATM in the context of gammaherpesvirus infection in vivo and support a model where ATM combines pro- and antiviral functions to facilitate both gammaherpesvirus-specific T cell immune response and viral reactivation in vivo. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. ATM/ATR-related checkpoint signals mediate arsenite-induced G{sub 2}/M arrest in primary aortic endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Tsou, Tsui-Chun; Tsai, Feng-Yuan; Yeh, Szu-Ching; Chang, Louis W. [National Health Research Institutes, Division of Environmental Health and Occupational Medicine, Miaoli County (Taiwan)

    2006-12-15

    Epidemiological studies have demonstrated a high association of inorganic arsenic exposure with vascular disease. Our recent in vitro studies have linked this vascular damage to vascular endothelial dysfunction induced by arsenic exposure. However, cell-cycle arrest induced by arsenic and its involvement in vascular dysfunction remain to be clarified. In this study, we employed primary porcine aortic endothelial cells to investigate regulatory mechanisms of G{sub 2}/M phase arrest induced by arsenite. Our study revealed that lower concentrations of arsenite (1 and 3 {mu}M) increased cell proliferation, whereas higher concentrations of arsenite (10, 20, and 30 {mu}M) inhibited cell proliferation together with correlated increases in G{sub 2}/M phase arrest. We found that this arsenite-induced G{sub 2}/M phase arrest was accompanied by accumulation and/or phosphorylation of checkpoint-related molecules, including p53, Cdc25B, Cdc25C, and securin. Inhibition of activations of these checkpoint-related molecules by caffeine significantly attenuated the 30-{mu}M arsenite-induced G{sub 2}/M phase arrest by 93%. Our data suggest that the DNA damage responsive kinases ATM (ataxia-telangiectasia mutated) and ATR (ATM and Rad3-related) play critical roles in arsenite-induced G{sub 2}/M phase arrest in aortic endothelial cells possibly via regulation of checkpoint-related signaling molecules including p53, Cdc25B, Cdc25C, and securin. (orig.)

  4. Integrating Gene Correction in the Reprogramming and Transdifferentiation Processes: A One-Step Strategy to Overcome Stem Cell-Based Gene Therapy Limitations

    Directory of Open Access Journals (Sweden)

    Seo-Young Lee

    2016-01-01

    Full Text Available The recent advent of induced pluripotent stem cells (iPSCs and gene therapy tools has raised the possibility of autologous cell therapy for rare genetic diseases. However, cellular reprogramming is inefficient in certain diseases such as ataxia telangiectasia, Fanconi anemia, LIG4 syndrome, and fibrodysplasia ossificans progressiva syndrome, owing to interference of the disease-related genes. To overcome these therapeutic limitations, it is necessary to fundamentally correct the abnormal gene during or prior to the reprogramming process. In addition, as genetic etiology of Parkinson’s disease, it has been well known that induced neural stem cells (iNSCs were progressively depleted by LRRK2 gene mutation, LRRK2 (G2019S. Thus, to maintain the induced NSCs directly derived from PD patient cells harboring LRRK2 (G2019S, it would be ideal to simultaneously treat the LRRK2 (G2019S fibroblast during the process of TD. Therefore, simultaneous reprogramming (or TD and gene therapy would provide the solution for therapeutic limitation caused by vulnerability of reprogramming or TD, in addition to being suitable for general application to the generation of autologous cell-therapy products for patients with genetic defects, thereby obviating the need for the arduous processes currently required.

  5. [Pathways for maintenance of mitochondrial DNA integrity and mitochondrial functions in cells exposed to ionizing radiation].

    Science.gov (United States)

    Gaziev, A I

    2013-01-01

    The analytical review deals with the results of studies devoted to mitochondrial DNA (mtDNA) disorders, the development of oxidative stress and possible pathways for the maintenance of mitochondrial functions in cells exposed to ionizing radiation (IR). Mitochondrial functions, which are closely related to the integrity of mtDNA, play a key role in many cellular processes. A wide range of degenerative diseases, carcinogenesis, and aging is associated with disturbances in mtDNA. MtDNA and the mitochondrion as a whole are increasingly considered as sensitive targets for cancer radio-chemotherapy. Knowledge of post-radiation processes in the mitochondria also facilitates creation of possible additional ways to reduce the radiation reaction of the organism. Injuries and mutations in mtDNA occur with a greater frequency than in the nuclear DNA (nDNA) in cells exposed to IR and other genotoxicants. On the other hand, functionally active copies of mtDNA can persist and survive in the cells exposed to clinically relevant doses of radiation. This safety is ensured by numerous copies of mtDNA in the cell, and due to their shielding from the effects of reactive oxygen (and nitrogen) species (ROS) by nucleoid proteins and by the operation of base excision repair in mitochondria. However, the generation of ROS increases in the mitochondria of cells exposed to IR. The increased generation of ROS in mitochondria can sometimes persist up to several days after the exposure of cells. The prolonged increased generation of ROS may be due to the involvement in the electron transport chain of the complexes of aberrant proteins expressed by the genes of mutated mtDNA copies. This may lead to the additional DNA damage, mitochondrial dysfunction, and instability of the nuclear genome. However, the development of oxidative stress can be restrained by antioxidant systems in the mitochondria. The key role here is played by activation of Mn-SOD2 and the protein p53. In addition, activation of

  6. Nature and role of high sister chromatid exchanges in Bloom syndrome cells. Some cytogenetic and immunological aspects.

    Science.gov (United States)

    Shiraishi, Y

    1990-12-01

    The phenomenon of sister chromatid exchange (SCE) is an interesting genetic event in metaphase chromosomes, even though its exact mechanism remains unknown. The fact that SCE can take place, whether "spontaneously" or induced by various agents, is in itself important, for such an event, involving damage and possible repair of bilateral loci in chromosomes, presents opportunities for modification of the chromosomal structure and/or function (e.g., oncogene activation). It has been assumed that under "normal" circumstances, SCE does not lead to any change in the functional genome, although this may not apply to abnormal conditions. The latter may be produced by a number of chemical agents (including various carcinogens) that lead to a significantly increased incidence of SCE in normal and malignant cells either in vitro or in vivo. In fact, SCE has been recognized and advocated as a most sensitive test for potentially mutagenic and/or carcinogenic agents. Thus, the broad field of SCE studies becomes of direct interest for and subject to exploration by those involved with cancer causation and biology. In this review, a synopsis of our experiences with high SCE mechanisms in Bloom syndrome cell lines will be presented, in connection with high SCE mutant cell line derived from ataxia telangiectasia (AT) and/or malignant transformation.

  7. Modulation of proteostasis counteracts oxidative stress and affects DNA base excision repair capacity in ATM-deficient cells.

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    Poletto, Mattia; Yang, Di; Fletcher, Sally C; Vendrell, Iolanda; Fischer, Roman; Legrand, Arnaud J; Dianov, Grigory L

    2017-09-29

    Ataxia telangiectasia (A-T) is a syndrome associated with loss of ATM protein function. Neurodegeneration and cancer predisposition, both hallmarks of A-T, are likely to emerge as a consequence of the persistent oxidative stress and DNA damage observed in this disease. Surprisingly however, despite these severe features, a lack of functional ATM is still compatible with early life, suggesting that adaptation mechanisms contributing to cell survival must be in place. Here we address this gap in our knowledge by analysing the process of human fibroblast adaptation to the lack of ATM. We identify profound rearrangement in cellular proteostasis occurring very early on after loss of ATM in order to counter protein damage originating from oxidative stress. Change in proteostasis, however, is not without repercussions. Modulating protein turnover in ATM-depleted cells also has an adverse effect on the DNA base excision repair pathway, the major DNA repair system that deals with oxidative DNA damage. As a consequence, the burden of unrepaired endogenous DNA lesions intensifies, progressively leading to genomic instability. Our study provides a glimpse at the cellular consequences of loss of ATM and highlights a previously overlooked role for proteostasis in maintaining cell survival in the absence of ATM function. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. Identification of gene-based responses in human blood cells exposed to alpha particle radiation.

    Science.gov (United States)

    Chauhan, Vinita; Howland, Matthew; Wilkins, Ruth

    2014-07-12

    The threat of a terrorist-precipitated nuclear event places humans at danger for radiological exposures. Isotopes which emit alpha (α)-particle radiation pose the highest risk. Currently, gene expression signatures are being developed for radiation biodosimetry and triage with respect to ionizing photon radiation. This study was designed to determine if similar gene expression profiles are obtained after exposures involving α-particles. Peripheral blood mononuclear cells (PBMCs) were used to identify sensitive and robust gene-based biomarkers of α-particle radiation exposure. Cells were isolated from healthy individuals and were irradiated at doses ranging from 0-1.5 Gy. Microarray technology was employed to identify transcripts that were differentially expressed relative to unirradiated cells 24 hours post-exposure. Statistical analysis identified modulated genes at each of the individual doses. Twenty-nine genes were common to all doses with expression levels ranging from 2-10 fold relative to control treatment group. This subset of genes was further assessed in independent complete white blood cell (WBC) populations exposed to either α-particles or X-rays using quantitative real-time PCR. This 29 gene panel was responsive in the α-particle exposed WBCs and was shown to exhibit differential fold-changes compared to X-irradiated cells, though no α-particle specific transcripts were identified. Current gene panels for photon radiation may also be applicable for use in α-particle radiation biodosimetry.

  9. Ceftaroline modulates the innate immune and host defense responses of immunocompetent cells exposed to cigarette smoke.

    Science.gov (United States)

    Bruno, A; Cipollina, C; Di Vincenzo, S; Siena, L; Dino, P; Di Gaudio, F; Gjomarkaj, M; Pace, E

    2017-09-05

    Cigarette smoke, the principal risk factor for chronic obstructive pulmonary disease (COPD), negatively influences the effectiveness of the immune system's response to a pathogen. The antibiotic ceftaroline exerts immune-modulatory effects in bronchial epithelial cells exposed to cigarette smoke. The present study aims to assess the effects of ceftaroline on TLR2 and TLR4 expression, LPS binding and TNF-α and human beta defensin (HBD2) release in an undifferentiated and PMA-differentiated human monocyte cell line (THP-1) exposed or not to cigarette smoke extracts (CSE). TLR2, TLR4, and LPS binding were assessed by flow cytometry, TNF-α and HBD2 release were evaluated by ELISA. The constitutive expression of TLR2 and TLR4 and LPS binding were higher in differentiated compared to undifferentiated THP-1 cells. In undifferentiated THP-1 cells, CSE increased TLR2 and TLR4 protein levels, LPS binding and TNF-α release and reduced HBD2 release and ceftaroline counteracted all these effects. In differentiated THP-1, CSE did not significantly affect TLR2 and TLR4 expression and LPS binding but reduced HBD2 release and increased TNF-α release. Ceftaroline counteracted the effects of CSE on HBD2 release in differentiated THP-1. Ceftaroline counteracts the effect of CSE in immune cells by increasing the effectiveness of the innate immune system. This effect may also assist in reducing pathogen activity and recurrent exacerbations in COPD patients. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Neutrophil capture by selectins on endothelial cells exposed to cigarette smoke.

    Science.gov (United States)

    Stone, P C W; Fisher, A C; Rainger, G E; Nash, G B

    2002-08-02

    We used a novel perfusion system to expose cultured human umbilical vein endothelial cells (HUVEC) to water-soluble components of cigarette smoke and study subsequent adhesion of flowing neutrophils. Neutrophils did not bind to HUVEC immediately after it had been exposed to cigarette smoke, but many adhered 90-150 min after exposure. The effect was reduced if the exposed medium was made serum-free, but this reduction was partially reversed if low density lipoprotein was added. Treatment of smoke-exposed HUVEC with antibodies against E-selectin or P-selectin reduced adhesion by approximately 50% or 75%, respectively; a combination of both antibodies essentially abolished adhesion. Enzyme-linked immunosorbent assay confirmed that exposure to smoke caused HUVEC to upregulate surface expression of E- and P-selectin. Thus, water-soluble constituent(s) of cigarette smoke cause efficient selectin-mediated capture of flowing neutrophils. This pro-inflammatory response may contribute to pathology associated with smoking, especially in tissues remote from the lung.

  11. Protective Effects of Hydroalcoholic Extract of Nasturtium officinale on Rat Blood Cells Exposed to Arsenic

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    Felor Zargari

    2015-06-01

    Full Text Available Background: Arsenic is one of the most toxic metalloids. Anemia and leukopenia are common results of poisoning with arsenic, which may happen due to a direct hemolytic or cytotoxic effect on blood cells. The aim of this study was to examine the effects of hydroalcoholic extract of Nasturtium officinale on blood cells and antioxidant enzymes in rats exposed to sodium (metaarsenite. Methods: 32 Male Sprague Dawley rats were randomly divided into four groups; Group I (normal healthy rats, Group II (treated with 5.5mg/kg of body weight of NaAsO2, Group III (treated with 500mg/kg of body weight of hydro-alcoholic extract of N. officinale, and Group IV (treated with group II and III supplementations. Blood samples were collected and red blood cell, white blood cell, hematocrit, hemoglobin, platelet, total protein and albumin levels and total antioxidant capacity were measured. Data was analyzed with Mann-Whitney U test. Results: WBC, RBC and Hct were decreased in the rats exposed to NaAsO2 (p<0.05. A significant increase was seen in RBC and Hct after treatment with the plant extract (p<0.05. There was no significant decrease in serum albumin and total protein in the groups exposed to NaAsO2 compared to the group I, but NaAsO2 decreased the total antioxidant capacity, significantly. Conclusion: The Nasturtium officinale extract have protective effect on arsenic-induced damage of blood cells.

  12. Cell surface differences of Naegleria fowleri and Naegleria lovaniensis exposed with surface markers.

    Science.gov (United States)

    González-Robles, Arturo; Castañón, Guadalupe; Cristóbal-Ramos, Ana Ruth; Hernández-Ramírez, Verónica Ivonne; Omaña-Molina, Maritza; Martínez-Palomo, Adolfo

    2007-12-01

    Differences in the distribution of diverse cell surface coat markers were found between Naegleria fowleri and Naegleria lovaniensis. The presence of carbohydrate-containing components in the cell coat of the two species was detected by selective staining with ruthenium red and alcian blue. Using both markers, N. fowleri presented a thicker deposit than N. lovaniensis. The existence of exposed mannose or glucose residues was revealed by discriminatory agglutination with the plant lectin Concanavalin A. These sugar residues were also visualized at the cell surface of these parasites either by transmission electron microscopy or by fluorescein-tagged Concanavalin A. Using this lectin cap formation was induced only in N. fowleri. The anionic sites on the cell surface detected by means of cationized ferritin were more apparent in N. fowleri. Biotinylation assays confirmed that even though the two amoebae species have some analogous plasma membrane proteins, there is a clear difference in their composition.

  13. Sirolimus Increases T-Cell Abundance in the Sun Exposed Skin of Kidney Transplant Recipients.

    Science.gov (United States)

    Burke, Michael Thomas; Sambira Nahum, Lauren C; Isbel, Nicole M; Carroll, Robert P; Soyer, Hans Peter; Francis, Ross; Bridge, Jennifer Anne; Hawley, Carmel; Oliver, Kimberly; Staatz, Christine E; Wells, James William

    2017-07-01

    Kidney transplant recipients (KTRs) receiving the mammalian target of rapamycin inhibitor sirolimus may display a reduced risk of skin cancer development compared to KTRs receiving calcineurin inhibitors. Despite studies investigating the effects of these 2 drug classes on T cells in patient blood, the effect these drugs may have in patient skin is not yet known. Fifteen patients with chronic kidney disease (not recipients of immunosuppressive drugs), and 30 KTRs (15 receiving a calcineurin inhibitor, and 15 receiving sirolimus) provided matched samples of blood, sun exposed (SE) and non-SE skin. The abundance of total CD8+ and CD4+ T cells, memory CD8+ and CD4+ T cells, and regulatory T (Treg) cells in each sample was then assessed by flow cytometry. Sirolimus treatment significantly increased absolute numbers of CD4+ T cells, memory CD8+- and CD4+ T cells, and Treg cells in SE skin versus paired samples of non-SE skin. No differences were found in the absolute number of any T cell subset in the blood. Correlation analysis revealed that the percentage of T cell subsets in the blood does not always accurately reflect the percentage of T-cell subsets in the skin of KTRs. Furthermore, sirolimus significantly disrupts the balance of memory CD4+ T cells in the skin after chronic sun exposure. This study demonstrated that immunosuppressive drug class and sun exposure modify the abundance of multiple T-cell subsets in the skin of KTRs. Correlation analysis revealed that the prevalence of Treg cells in KTR blood does not accurately reflect the prevalence of Treg cells in KTR skin.

  14. Phenotypic modifications in Staphylococcus aureus cells exposed to high concentrations of vancomycin and teicoplanin

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    Fábio D.A. Gonçalves

    2016-01-01

    Full Text Available Bacterial cells are known to change the fatty acid composition of the phospholipids as a phenotypic response to environmental conditions and to the presence of toxic compounds such as antibiotics. In the present study, Staphylococcus aureus cells collected during the exponential growth phase were challenged with 50 and 100 mg/L of vancomycin and teicoplanin, which are concentrations high enough to kill the large majority of the cell population. Colony-forming unit counts showed biphasic killing kinetics, typical for persister cell enrichment, in both antibiotics and concentrations tested. However, fluorescence microscopy showed the existence of viable but non-culturable (VBNC cells in a larger number than that of possible persister cells.The analysis of the fatty acid composition of the cells showed that, following antibiotic exposure up to 6 h, the survivor cells have an increased percentage of saturated fatty acids, a significant reduced percentage of branched fatty acids and an increased iso/anteiso branched fatty acid ratio when compared to cells exhibiting a regular phenotype. This should result in lower membrane fluidity. However, cells exposed for 8-24 h presented an increased branched/saturated and lower iso/anteiso branched fatty acid ratios, and thus increased membrane fluidity. Furthermore, the phenotypic changes were transmitted to daughter cells grown in drug-free media. The fact that VBNC cells presented nearly the same fatty acid composition as those obtained after cell growth in drug-free media, which could only be the result of growth of persister cells, suggest that VBNC and persister phenotypes share the same type of response to antibiotics at the lipid level.

  15. Assessment of DNA integrity (COMET assay) in sperm cells of boron-exposed workers.

    Science.gov (United States)

    Duydu, Yalçin; Başaran, Nurşen; Ustündağ, Aylin; Aydin, Sevtap; Undeğer, Ulkü; Ataman, Osman Yavuz; Aydos, Kaan; Düker, Yalçin; Ickstadt, Katja; Waltrup, Britta Schulze; Golka, Klaus; Bolt, Hermann M

    2012-01-01

    An extension of a male reproductive study conducted in a boric acid/borate production zone at Bandırma, Turkey, is presented. The relation between DNA-strand breaks (COMET assay, neutral and alkaline version) in sperm cells and previously described sperm quality parameters was investigated in boron-exposed males. A correlation between blood boron levels and mean DNA-strand breaks in sperm was weak, and DNA-strand breaks in sperm were statistically not different between control and exposed groups. Therefore, increasing boron exposures had no additional contribution in addition to already pre-existing DNA-strand breaks in the sperm cells. Weak but statistically significant correlations between DNA-strand breaks and motility/morphology parameters of sperm samples were observed in the neutral version of the COMET assay, while correlations between the same variables were statistically not significant in the alkaline version. A likely reason for these negative results, even in highly exposed humans, is that experimental exposures that had led to reproductive toxicity in animals were significantly higher than any boron exposures, which may be reached under realistic human conditions.

  16. A genome-wide analysis of gene-caffeine consumption interaction on basal cell carcinoma.

    Science.gov (United States)

    Li, Xin; Cornelis, Marilyn C; Liang, Liming; Song, Fengju; De Vivo, Immaculata; Giovannucci, Edward; Tang, Jean Y; Han, Jiali

    2016-12-01

    Animal models have suggested that oral or topical administration of caffeine could inhibit ultraviolet-induced carcinogenesis via the ataxia telangiectasia and rad3 (ATR)-related apoptosis. Previous epidemiological studies have demonstrated that increased caffeine consumption is associated with reduced risk of basal cell carcinoma (BCC). To identify common genetic markers that may modify this association, we tested gene-caffeine intake interaction on BCC risk in a genome-wide analysis. We included 3383 BCC cases and 8528 controls of European ancestry from the Nurses' Health Study and Health Professionals Follow-up Study. Single nucleotide polymorphism (SNP) rs142310826 near the NEIL3 gene showed a genome-wide significant interaction with caffeine consumption (P = 1.78 × 10-8 for interaction) on BCC risk. There was no gender difference for this interaction (P = 0.64 for heterogeneity). NEIL3, a gene belonging to the base excision DNA repair pathway, encodes a DNA glycosylase that recognizes and removes lesions produced by oxidative stress. In addition, we identified several loci with P value for interaction caffeine consumption-related SNPs reported by previous genome-wide association studies and risk of BCC, both individually and jointly, but found no significant association. In sum, we identified a DNA repair gene that could be involved in caffeine-mediated skin tumor inhibition. Further studies are warranted to confirm these findings. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  17. Diagnostic and pathogenetic role of café-au-lait macules in nevoid basal cell carcinoma syndrome

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    Ponti Giovanni

    2012-10-01

    Full Text Available Abstract Café au lait spots (CALS are common dermatologic findings that can at the same time arise in a variety of pathologic conditions such as Neurofibromatosis type 1 (NF1, together with numerous hereditary syndromes for which they represent either diagnostic criteria or associated elements (McCune Albright, Silver-Russell, LEOPARD, Ataxia-Telangiectasia. A review of the literature also revealed two cases of association with NBCCS. We report here the case of a female proband with CALS associated to Nevoid Basal Cell Carcinoma Syndrome (NBCCS with known PTCH1 germline mutation (C.1348-2A>G who had been misdiagnosed with NF1 in her childhood because of 5 CALS and cutaneous nodules. The patient presented a giant cell tumor of the skin, palmar and calcaneal epidermoidal cystic nodules, odontogenic keratocystic tumors and deformity of the jaw profile. Her family history brought both her brother and father to our attention because of the presence of KCOTs diagnosed at early age: after genetic testing, the same PTCH1 germline mutation was identified in the three family members. Clinical criteria are used for discerning NF1 diagnosis (size, number and onset age, while there are no definite guidelines concerning CALS except for their presence. In our experience, we have noted an association of CALS with NBCCS; this seems interesting because we already know clinical criteria are a dynamic entity and can be modified by epidemiologic evidences.

  18. Analysis of hypermethylation and expression profiles of APC and ATM genes in patients with oral squamous cell carcinoma

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    Rigi-Ladiz Mohammad

    2011-11-01

    Full Text Available Abstract Background Adenomatous polyposis coli (APC and Ataxia-telangiectasia-mutated (ATM gene products have an important role in cell cycle control and maintenance of genomic stability. Our aim was to analyze ATM and APC methylation and its relationship with oral squamous cell carcinoma (OSCC. Materials and methods Eighty-four OSCC tissues that have been fixed in paraffin along with 57 control oral samples have been used for analyzing promoter methylation of ATM and APC genes by Methylation Specific Polymerase Chain Reaction (MS-PCR. In addition, 10 cases of OSCC and the same of matched controls were examined for estimating expression of the above mentioned genes using Real-Time Reverse-Transcription PCR. Results Observed promoter methylations were 71.42% and 87.71% for the APC gene and 88.09% and 77.19% for the ATM gene in cases and controls, respectively. Analysis of these data showed that promoter methylation at APC was significantly different in cases compared to healthy controls (p = 0.01, but no difference was detected for the ATM gene. Furthermore, the mRNA expression levels did not differ statistically between cases and controls for both ATM (cases = 9, controls = 10 and APC (cases = 11, controls = 10 genes. Conclusions Our results, for the first time, provide methylation profiles of ATM and APC genes in a sample of patients with OSCC in a southeast Iranian population. The present data support related evidence of APC methylation effect on OSCC development.

  19. Diagnostic and pathogenetic role of café-au-lait macules in nevoid basal cell carcinoma syndrome.

    Science.gov (United States)

    Ponti, Giovanni; Tomasi, Aldo; Pastorino, Lorenza; Ruini, Cristel; Guarneri, Carmelo; Mandel, Victor Desmond; Seidenari, Stefania; Pellacani, Giovanni

    2012-10-29

    Café au lait spots (CALS) are common dermatologic findings that can at the same time arise in a variety of pathologic conditions such as Neurofibromatosis type 1 (NF1), together with numerous hereditary syndromes for which they represent either diagnostic criteria or associated elements (McCune Albright, Silver-Russell, LEOPARD, Ataxia-Telangiectasia). A review of the literature also revealed two cases of association with NBCCS. We report here the case of a female proband with CALS associated to Nevoid Basal Cell Carcinoma Syndrome (NBCCS) with known PTCH1 germline mutation (C.1348-2A>G) who had been misdiagnosed with NF1 in her childhood because of 5 CALS and cutaneous nodules. The patient presented a giant cell tumor of the skin, palmar and calcaneal epidermoidal cystic nodules, odontogenic keratocystic tumors and deformity of the jaw profile. Her family history brought both her brother and father to our attention because of the presence of KCOTs diagnosed at early age: after genetic testing, the same PTCH1 germline mutation was identified in the three family members. Clinical criteria are used for discerning NF1 diagnosis (size, number and onset age), while there are no definite guidelines concerning CALS except for their presence. In our experience, we have noted an association of CALS with NBCCS; this seems interesting because we already know clinical criteria are a dynamic entity and can be modified by epidemiologic evidences.

  20. Single Walled Carbon Nanotubes Exhibit Dual-Phase Regulation to Exposed Arabidopsis Mesophyll Cells

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    Huang Peng

    2011-01-01

    Full Text Available Abstract Herein we are the first to report that single-walled carbon nanotubes (SWCNTs exhibit dual-phase regulation to Arabidopsis mesophyll cells exposed to different concentration of SWCNTs. The mesophyll protoplasts were prepared by enzyme digestion, and incubated with 15, 25, 50, 100 μg/ml SWCNTs for 48 h, and then were observed by optical microscopy and transmission electron microscopy, the reactive oxygen species (ROS generation was measured. Partial protoplasts were stained with propidium iodide and 4'-6- diamidino-2-phenylindole, partial protoplasts were incubated with fluorescein isothiocyanate-labeled SWCNTs, and observed by fluorescence microscopy. Results showed that SWCNTs could traverse both the plant cell wall and cell membrane, with less than or equal to 50 μg/ml in the culture medium, SWCNTs stimulated plant cells to grow out trichome clusters on their surface, with more than 50 μg/ml SWCNTs in the culture medium, SWCNTs exhibited obvious toxic effects to the protoplasts such as increasing generation of ROS, inducing changes of protoplast morphology, changing green leaves into yellow, and inducing protoplast cells' necrosis and apoptosis. In conclusion, single walled carbon nanotubes can get through Arabidopsis mesophyll cell wall and membrane, and exhibit dose-dependent dual-phase regulation to Arabidopsis mesophyll protoplasts such as low dose stimulating cell growth, and high dose inducing cells' ROS generation, necrosis or apoptosis.

  1. Spinocerebellar ataxia: miRNAs expose biological pathways underlying pervasive Purkinje cell degeneration.

    Science.gov (United States)

    van der Stijl, Rogier; Withoff, Sebo; Verbeek, Dineke S

    2017-12-01

    Recent work has demonstrated the importance of miRNAs in the pathogenesis of various brain disorders including the neurodegenerative disorder spinocerebellar ataxia (SCA). This review focuses on the role of miRNAs in the shared pathogenesis of the different SCA types. We examine the novel findings of a recent cell-type-specific RNA-sequencing study in mouse brain and discuss how the identification of Purkinje-cell-enriched miRNAs highlights biological pathways that expose the mechanisms behind pervasive Purkinje cell degeneration in SCA. These key pathways are likely to contain targets for therapeutic development and represent potential candidate genes for genetically unsolved SCAs. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Colon stem cell and crypt dynamics exposed by cell lineage reconstruction.

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    Yitzhak Reizel

    2011-07-01

    Full Text Available Stem cell dynamics in vivo are often being studied by lineage tracing methods. Our laboratory has previously developed a retrospective method for reconstructing cell lineage trees from somatic mutations accumulated in microsatellites. This method was applied here to explore different aspects of stem cell dynamics in the mouse colon without the use of stem cell markers. We first demonstrated the reliability of our method for the study of stem cells by confirming previously established facts, and then we addressed open questions. Our findings confirmed that colon crypts are monoclonal and that, throughout adulthood, the process of monoclonal conversion plays a major role in the maintenance of crypts. The absence of immortal strand mechanism in crypts stem cells was validated by the age-dependent accumulation of microsatellite mutations. In addition, we confirmed the positive correlation between physical and lineage proximity of crypts, by showing that the colon is separated into small domains that share a common ancestor. We gained new data demonstrating that colon epithelium is clustered separately from hematopoietic and other cell types, indicating that the colon is constituted of few progenitors and ruling out significant renewal of colonic epithelium from hematopoietic cells during adulthood. Overall, our study demonstrates the reliability of cell lineage reconstruction for the study of stem cell dynamics, and it further addresses open questions in colon stem cells. In addition, this method can be applied to study stem cell dynamics in other systems.

  3. Microtubule Formation and Activities of Antioxidative Enzymes in PC12 Cells Exposed to Phosphatidylcholine Hydroperoxides

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    Yukako Yamanaka

    2012-11-01

    Full Text Available Aging increases free radical generation and lipid oxidation and, thereby, mediates neurodegenerative diseases. As the brain is rich in lipids (polyunsaturated fatty acids, the antioxidative system plays an important role in protecting brain tissues from oxidative injury. The changes in microtubule formation and antioxidative enzyme activities have been investigated in rat pheochromocytoma PC12 cells exposed to various concentrations of phosphatidylcholine hydroperoxides (PCOOH. We measured three typical antioxidative enzymes, superoxide dismutase (SOD, glutathione peroxidase (GPx, and catalase (CAT. The microtubule assembly system was dependent on the antioxidative enzyme system in cells exposed to oxidative stress. The activities of the three enzymes increased in a PCOOH exposure-dependent manner. In particular, the changes in the activity as a result of PCOOH exposure were similar in the three antioxidative enzymes. This is the first report indicating the compatibility between the tubulin-microtubule and antioxidative enzyme systems in cells that deteriorate as a result of phospholipid hydroperoxide administration from an exterior source. The descending order of sensitivity of the three enzymes to PCOOH is also discussed.

  4. Cisplatin resistance in non-small cell lung cancer cells is associated with an abrogation of cisplatin-induced G2/M cell cycle arrest.

    Science.gov (United States)

    Sarin, Navin; Engel, Florian; Kalayda, Ganna V; Mannewitz, Mareike; Cinatl, Jindrich; Rothweiler, Florian; Michaelis, Martin; Saafan, Hisham; Ritter, Christoph A; Jaehde, Ulrich; Frötschl, Roland

    2017-01-01

    The efficacy of cisplatin-based chemotherapy in cancer is limited by the occurrence of innate and acquired drug resistance. In order to better understand the mechanisms underlying acquired cisplatin resistance, we have compared the adenocarcinoma-derived non-small cell lung cancer (NSCLC) cell line A549 and its cisplatin-resistant sub-line A549rCDDP2000 with regard to cisplatin resistance mechanisms including cellular platinum accumulation, DNA-adduct formation, cell cycle alterations, apoptosis induction and activation of key players of DNA damage response. In A549rCDDP2000 cells, a cisplatin-induced G2/M cell cycle arrest was lacking and apoptosis was reduced compared to A549 cells, although equitoxic cisplatin concentrations resulted in comparable platinum-DNA adduct levels. These differences were accompanied by changes in the expression of proteins involved in DNA damage response. In A549 cells, cisplatin exposure led to a significantly higher expression of genes coding for proteins mediating G2/M arrest and apoptosis (mouse double minute 2 homolog (MDM2), xeroderma pigmentosum complementation group C (XPC), stress inducible protein (SIP) and p21) compared to resistant cells. This was underlined by significantly higher protein levels of phosphorylated Ataxia telangiectasia mutated (pAtm) and p53 in A549 cells compared to their respective untreated control. The results were compiled in a preliminary model of resistance-associated signaling alterations. In conclusion, these findings suggest that acquired resistance of NSCLC cells against cisplatin is the consequence of altered signaling leading to reduced G2/M cell cycle arrest and apoptosis.

  5. Cisplatin resistance in non-small cell lung cancer cells is associated with an abrogation of cisplatin-induced G2/M cell cycle arrest.

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    Navin Sarin

    Full Text Available The efficacy of cisplatin-based chemotherapy in cancer is limited by the occurrence of innate and acquired drug resistance. In order to better understand the mechanisms underlying acquired cisplatin resistance, we have compared the adenocarcinoma-derived non-small cell lung cancer (NSCLC cell line A549 and its cisplatin-resistant sub-line A549rCDDP2000 with regard to cisplatin resistance mechanisms including cellular platinum accumulation, DNA-adduct formation, cell cycle alterations, apoptosis induction and activation of key players of DNA damage response. In A549rCDDP2000 cells, a cisplatin-induced G2/M cell cycle arrest was lacking and apoptosis was reduced compared to A549 cells, although equitoxic cisplatin concentrations resulted in comparable platinum-DNA adduct levels. These differences were accompanied by changes in the expression of proteins involved in DNA damage response. In A549 cells, cisplatin exposure led to a significantly higher expression of genes coding for proteins mediating G2/M arrest and apoptosis (mouse double minute 2 homolog (MDM2, xeroderma pigmentosum complementation group C (XPC, stress inducible protein (SIP and p21 compared to resistant cells. This was underlined by significantly higher protein levels of phosphorylated Ataxia telangiectasia mutated (pAtm and p53 in A549 cells compared to their respective untreated control. The results were compiled in a preliminary model of resistance-associated signaling alterations. In conclusion, these findings suggest that acquired resistance of NSCLC cells against cisplatin is the consequence of altered signaling leading to reduced G2/M cell cycle arrest and apoptosis.

  6. [The intervention of nicotinamide on skin melanocyte's cell proliferation after UVA (365 nm) exposed.].

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    Patam, Muhammad; Jin, Xi-peng; Pan, Jian-ying; Shen, Guang-zu; Jin, Tai-Yi

    2005-02-01

    To investigate the interference effect of nicotinamide on UVA-induced cell proliferation in human skin melanocyte. To apply the optimum UVA dose expected to cause cell proliferation: 0.2 cm2, nicotinamide was added after the 0.2 cm2 UVA exposure immediately or 48 h later, then the rate of cell proliferation, calcium concentration and the activities of Na+-K+, Ca2+-ATP enzymes of melanocytes were measured respectively. After treatment with 1.000 mg/ml nicotinamide following UVA exposure, the rate of cell proliferation was decreased significantly 24 hours later. Treatment with 0.125 mg/ml nicotinamide 48 hours after UVA exposure also significantly inhibited the cell proliferation; 1.25 mg/ml nicotinamide increased calcium concentration in cells; 0.250 mg/ml nicotinamide increased the activities of Na+-K+, Ca2+-ATP enzymes in melanocytes (P Nicotinamide has more obvious effect on inhibiting melanocyte's proliferation if added immediately following UVA exposure. Our discovery indicated that nicotinamide may affect the melanocyte through modulating the calcium concentration. It is possible to consider nicotinamide as an efficient and safe sun screen to provide a certain level of protection for UVA exposed skin.

  7. Evaluation of cell types for assessment of cytogenetic damage in arsenic exposed population

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    Singh Keshav K

    2008-05-01

    Full Text Available Abstract Background Cytogenetic biomarkers are essential for assessing environmental exposure, and reflect adverse human health effects such as cellular damage. Arsenic is a potential clastogen and aneugen. In general, the majority of the studies on clastogenic effects of arsenic are based on frequency of micronuclei (MN study in peripheral lymphocytes, urothelial and oral epithelial cells. To find out the most suitable cell type, here, we compared cytogenetic damage through MN assay in (a various populations exposed to arsenic through drinking water retrieved from literature review, as also (b arsenic-induced Bowen's patients from our own survey. Results For literature review, we have searched the Pubmed database for English language journal articles using the following keywords: "arsenic", "micronuclei", "drinking water", and "human" in various combinations. We have selected 13 studies consistent with our inclusion criteria that measured micronuclei in either one or more of the above-mentioned three cell types, in human samples. Compared to urothelial and buccal mucosa cells, the median effect sizes measured by the difference between people with exposed and unexposed, lymphocyte based MN counts were found to be stronger. This general pattern pooled from 10 studies was consistent with our own set of three earlier studies. MN counts were also found to be stronger for lymphocytes even in arsenic-induced Bowen's patients (cases compared to control individuals having arsenic-induced non-cancerous skin lesions. Conclusion Overall, it can be concluded that MN in lymphocytes may be superior to other epithelial cells for studying arsenic-induced cytogenetic damage.

  8. Altered global gene expression profiles in human gastrointestinal epithelial Caco2 cells exposed to nanosilver

    Directory of Open Access Journals (Sweden)

    Saura C. Sahu

    2016-01-01

    Full Text Available Extensive consumer exposure to food- and cosmetics-related consumer products containing nanosilver is of public safety concern. Therefore, there is a need for suitable in vitro models and sensitive predictive rapid screening methods to assess their toxicity. Toxicogenomic profile showing subtle changes in gene expressions following nanosilver exposure is a sensitive toxicological endpoint for this purpose. We evaluated the Caco2 cells and global gene expression profiles as tools for predictive rapid toxicity screening of nanosilver. We evaluated and compared the gene expression profiles of Caco-2 cells exposed to 20 nm and 50 nm nanosilver at a concentration 2.5 μg/ml. The global gene expression analysis of Caco2 cells exposed to 20 nm nanosilver showed that a total of 93 genes were altered at 4 h exposure, out of which 90 genes were up-regulated and 3 genes were down-regulated. The 24 h exposure of 20 nm silver altered 15 genes in Caco2 cells, out of which 14 were up-regulated and one was down-regulated. The most pronounced changes in gene expression were detected at 4 h. The greater size (50 nm nanosilver at 4 h exposure altered more genes by more different pathways than the smaller (20 nm one. Metallothioneins and heat shock proteins were highly up-regulated as a result of exposure to both the nanosilvers. The cellular pathways affected by the nanosilver exposure is likely to lead to increased toxicity. The results of our study presented here suggest that the toxicogenomic characterization of Caco2 cells is a valuable in vitro tool for assessing toxicity of nanomaterials such as nanosilver.

  9. Purkinje Cell Degeneration in pcd Mice Reveals Large Scale Chromatin Reorganization and Gene Silencing Linked to Defective DNA Repair*

    Science.gov (United States)

    Baltanás, Fernando C.; Casafont, Iñigo; Lafarga, Vanesa; Weruaga, Eduardo; Alonso, José R.; Berciano, María T.; Lafarga, Miguel

    2011-01-01

    DNA repair protects neurons against spontaneous or disease-associated DNA damage. Dysfunctions of this mechanism underlie a growing list of neurodegenerative disorders. The Purkinje cell (PC) degeneration mutation causes the loss of nna1 expression and is associated with the postnatal degeneration of PCs. This PC degeneration dramatically affects nuclear architecture and provides an excellent model to elucidate the nuclear mechanisms involved in a whole array of neurodegenerative disorders. We used immunocytochemistry for histone variants and components of the DNA damage response, an in situ transcription assay, and in situ hybridization for telomeres to analyze changes in chromatin architecture and function. We demonstrate that the phosphorylation of H2AX, a DNA damage signal, and the trimethylation of the histone H4K20, a repressive mark, in extensive domains of genome are epigenetic hallmarks of chromatin in degenerating PCs. These histone modifications are associated with a large scale reorganization of chromatin, telomere clustering, and heterochromatin-induced gene silencing, all of them key factors in PC degeneration. Furthermore, ataxia telangiectasia mutated and 53BP1, two components of the DNA repair pathway, fail to be concentrated in the damaged chromatin compartments, even though the expression levels of their coding genes were slightly up-regulated. Although the mechanism by which Nna1 loss of function leads to PC neurodegeneration is undefined, the progressive accumulation of DNA damage in chromosome territories irreversibly compromises global gene transcription and seems to trigger PC degeneration and death. PMID:21700704

  10. Purkinje cell degeneration in pcd mice reveals large scale chromatin reorganization and gene silencing linked to defective DNA repair.

    Science.gov (United States)

    Baltanás, Fernando C; Casafont, Iñigo; Lafarga, Vanesa; Weruaga, Eduardo; Alonso, José R; Berciano, María T; Lafarga, Miguel

    2011-08-12

    DNA repair protects neurons against spontaneous or disease-associated DNA damage. Dysfunctions of this mechanism underlie a growing list of neurodegenerative disorders. The Purkinje cell (PC) degeneration mutation causes the loss of nna1 expression and is associated with the postnatal degeneration of PCs. This PC degeneration dramatically affects nuclear architecture and provides an excellent model to elucidate the nuclear mechanisms involved in a whole array of neurodegenerative disorders. We used immunocytochemistry for histone variants and components of the DNA damage response, an in situ transcription assay, and in situ hybridization for telomeres to analyze changes in chromatin architecture and function. We demonstrate that the phosphorylation of H2AX, a DNA damage signal, and the trimethylation of the histone H4K20, a repressive mark, in extensive domains of genome are epigenetic hallmarks of chromatin in degenerating PCs. These histone modifications are associated with a large scale reorganization of chromatin, telomere clustering, and heterochromatin-induced gene silencing, all of them key factors in PC degeneration. Furthermore, ataxia telangiectasia mutated and 53BP1, two components of the DNA repair pathway, fail to be concentrated in the damaged chromatin compartments, even though the expression levels of their coding genes were slightly up-regulated. Although the mechanism by which Nna1 loss of function leads to PC neurodegeneration is undefined, the progressive accumulation of DNA damage in chromosome territories irreversibly compromises global gene transcription and seems to trigger PC degeneration and death.

  11. Germline Mutations of the Ataxia-Telangiectasia Gene, ATM, as a Risk Factor for Radiation-Associated Breast Cancer

    National Research Council Canada - National Science Library

    Offit, Kenneth

    1998-01-01

    ... of breast cancer after exposure to a defined dose of therapeutic irradiation. The study population will be comprised of women who have developed breast cancer after treatment for Hodgkin's Disease...

  12. Screening for Ataxia-Telangiectasia Mutations in a Population-Based Sample of Women with Early-Onset Breast Cancer

    Science.gov (United States)

    1999-09-01

    G, Itnyre J (1997) Prevalence and contribution of BRCA1 mutations in breast and ovarian cancer: results from three U.S. population-based case-control...GTTAAGAA two of these cases, AT1ILA and AT119LA, the families were consanguineous , and the mutations were homo- zygous, simplifying the interpretation...screening efficiency and expand significantly the number of cases to be screened. Assessment of the prevalence of ATM mutations in breast cancer

  13. Innate Lymphoid Cells Mediate Pulmonary Eosinophilic Inflammation, Airway Mucous Cell Metaplasia, and Type 2 Immunity in Mice Exposed to Ozone.

    Science.gov (United States)

    Kumagai, Kazuyoshi; Lewandowski, Ryan P; Jackson-Humbles, Daven N; Buglak, Nicholas; Li, Ning; White, Kaylin; Van Dyken, Steven J; Wagner, James G; Harkema, Jack R

    2017-08-01

    Exposure to elevated levels of ambient ozone in photochemical smog is associated with eosinophilic airway inflammation and nonatopic asthma in children. In the present study, we determined the role of innate lymphoid cells (ILCs) in the pathogenesis of ozone-induced nonatopic asthma by using lymphoid cell-sufficient C57BL/6 mice, ILC-sufficient Rag2 -/- mice (devoid of T and B cells), and ILC-deficient Rag2 -/- Il2rg -/- mice (depleted of all lymphoid cells including ILCs). Mice were exposed to 0 or 0.8 parts per million ozone for 1 day or 9 consecutive weekdays (4 hr/day). A single exposure to ozone caused neutrophilic inflammation, airway epithelial injury, and reparative DNA synthesis in all strains of mice, irrespective of the presence or absence of ILCs. In contrast, 9-day exposures induced eosinophilic inflammation and mucous cell metaplasia only in the lungs of ILC-sufficient mice. Repeated ozone exposures also elicited increased messenger RNA expression of transcripts associated with type 2 immunity and airway mucus production in ILC-sufficient mice. ILC-deficient mice repeatedly exposed to ozone had no pulmonary pathology or increased gene expression related to type 2 immunity. These results suggest a new paradigm for the biologic mechanisms underlying the development of a phenotype of childhood nonatopic asthma that has been linked to ambient ozone exposures.

  14. Radioprotection and Cell Cycle Arrest of Intestinal Epithelial Cells by Darinaparsin, a Tumor Radiosensitizer

    Energy Technology Data Exchange (ETDEWEB)

    Tian, Junqiang; Doi, Hiroshi [Department of Radiation Oncology, School of Medicine, Stanford University, Stanford, California (United States); Saar, Matthias; Santos, Jennifer [Department of Urology, School of Medicine, Stanford University, Stanford, California (United States); Li, Xuejun; Peehl, Donna M. [Department of Radiation Oncology, School of Medicine, Stanford University, Stanford, California (United States); Knox, Susan J., E-mail: sknox@stanford.edu [Department of Radiation Oncology, School of Medicine, Stanford University, Stanford, California (United States)

    2013-12-01

    Purpose: It was recently reported that the organic arsenic compound darinaparsin (DPS) is a cytotoxin and radiosensitizer of tumor cells in vitro and in subcutaneous xenograft tumors. Surprisingly, it was also found that DPS protects normal intestinal crypt epithelial cells (CECs) from clonogenic death after ionizing radiation (IR). Here we tested the DPS radiosensitizing effect in a clinically relevant model of prostate cancer and explored the radioprotective effect and mechanism of DPS on CECs. Methods and Materials: The radiation modification effect of DPS was tested in a mouse model of orthotopic xenograft prostate cancer and of IR-induced acute gastrointestinal syndrome. The effect of DPS on CEC DNA damage and DNA damage responses was determined by immunohistochemistry. Results: In the mouse model of IR-induced gastrointestinal syndrome, DPS treatment before IR accelerated recovery from body weight loss and increased animal survival. DPS decreased post-IR DNA damage and cell death, suggesting that the radioprotective effect was mediated by enhanced DNA damage repair. Shortly after DPS injection, significant cell cycle arrest was observed in CECs at both G1/S and G2/M checkpoints, which was accompanied by the activation of cell cycle inhibitors p21 and growth arrest and DNA-damage-inducible protein 45 alpha (GADD45A). Further investigation revealed that DPS activated ataxia telangiectasia mutated (ATM), an important inducer of DNA damage repair and cell cycle arrest. Conclusions: DPS selectively radioprotected normal intestinal CECs and sensitized prostate cancer cells in a clinically relevant model. This effect may be, at least in part, mediated by DNA damage response activation and has the potential to significantly increase the therapeutic index of radiation therapy.

  15. Radioprotection and cell cycle arrest of intestinal epithelial cells by darinaparsin, a tumor radiosensitizer.

    Science.gov (United States)

    Tian, Junqiang; Doi, Hiroshi; Saar, Matthias; Santos, Jennifer; Li, Xuejun; Peehl, Donna M; Knox, Susan J

    2013-12-01

    It was recently reported that the organic arsenic compound darinaparsin (DPS) is a cytotoxin and radiosensitizer of tumor cells in vitro and in subcutaneous xenograft tumors. Surprisingly, it was also found that DPS protects normal intestinal crypt epithelial cells (CECs) from clonogenic death after ionizing radiation (IR). Here we tested the DPS radiosensitizing effect in a clinically relevant model of prostate cancer and explored the radioprotective effect and mechanism of DPS on CECs. The radiation modification effect of DPS was tested in a mouse model of orthotopic xenograft prostate cancer and of IR-induced acute gastrointestinal syndrome. The effect of DPS on CEC DNA damage and DNA damage responses was determined by immunohistochemistry. In the mouse model of IR-induced gastrointestinal syndrome, DPS treatment before IR accelerated recovery from body weight loss and increased animal survival. DPS decreased post-IR DNA damage and cell death, suggesting that the radioprotective effect was mediated by enhanced DNA damage repair. Shortly after DPS injection, significant cell cycle arrest was observed in CECs at both G1/S and G2/M checkpoints, which was accompanied by the activation of cell cycle inhibitors p21 and growth arrest and DNA-damage-inducible protein 45 alpha (GADD45A). Further investigation revealed that DPS activated ataxia telangiectasia mutated (ATM), an important inducer of DNA damage repair and cell cycle arrest. DPS selectively radioprotected normal intestinal CECs and sensitized prostate cancer cells in a clinically relevant model. This effect may be, at least in part, mediated by DNA damage response activation and has the potential to significantly increase the therapeutic index of radiation therapy. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. T Cell Activation in South African HIV-Exposed Infants Correlates with Ochratoxin A Exposure

    Directory of Open Access Journals (Sweden)

    Lianna Frances Wood

    2017-12-01

    Full Text Available The introduction of non-breastmilk foods to HIV-infected infants is associated with increased levels of immune activation, which can impact the rate of HIV disease progression. This is particularly relevant in countries where mother-to-child transmission of HIV still occurs at unacceptable levels. The goal of this study was to evaluate the levels of the toxic food contaminant ochratoxin A (OTA in HIV-exposed South African infants that are either breastfed or consuming non-breast milk foods. OTA is a common mycotoxin, found in grains and soil, which is toxic at high doses but has immunomodulatory properties at lower doses. Samples from HIV-exposed and HIV-unexposed infants enrolled in prospective observational cohort studies were collected and analyzed at birth through 14 weeks of age. We observed that infants consuming non-breast milk foods had significantly higher plasma levels of OTA at 6 weeks of age compared to breastfed infants, increasing until 8 weeks of age. The blood levels of OTA detected were comparable to levels observed in OTA-endemic communities. OTA plasma levels correlated with HIV target cell activation (CCR5 and HLADR expression on CD4+ T cells and plasma levels of the inflammatory cytokine CXCL10. These findings provide evidence that elevated OTA levels in South African infants are associated with the consumption of non-breastmilk foods and activation of the immune system. Reducing infant OTA exposure has the potential to reduce immune activation and provide health benefits, particularly in those infants who are HIV-exposed or HIV-infected.

  17. T Cell Activation in South African HIV-Exposed Infants Correlates with Ochratoxin A Exposure.

    Science.gov (United States)

    Wood, Lianna Frances; Wood, Matthew P; Fisher, Bridget S; Jaspan, Heather B; Sodora, Donald L

    2017-01-01

    The introduction of non-breastmilk foods to HIV-infected infants is associated with increased levels of immune activation, which can impact the rate of HIV disease progression. This is particularly relevant in countries where mother-to-child transmission of HIV still occurs at unacceptable levels. The goal of this study was to evaluate the levels of the toxic food contaminant ochratoxin A (OTA) in HIV-exposed South African infants that are either breastfed or consuming non-breast milk foods. OTA is a common mycotoxin, found in grains and soil, which is toxic at high doses but has immunomodulatory properties at lower doses. Samples from HIV-exposed and HIV-unexposed infants enrolled in prospective observational cohort studies were collected and analyzed at birth through 14 weeks of age. We observed that infants consuming non-breast milk foods had significantly higher plasma levels of OTA at 6 weeks of age compared to breastfed infants, increasing until 8 weeks of age. The blood levels of OTA detected were comparable to levels observed in OTA-endemic communities. OTA plasma levels correlated with HIV target cell activation (CCR5 and HLADR expression on CD4+ T cells) and plasma levels of the inflammatory cytokine CXCL10. These findings provide evidence that elevated OTA levels in South African infants are associated with the consumption of non-breastmilk foods and activation of the immune system. Reducing infant OTA exposure has the potential to reduce immune activation and provide health benefits, particularly in those infants who are HIV-exposed or HIV-infected.

  18. Regulation of T Helper Cell Responses During Antigen Presentation by Norepinephrine-exposed Endothelial Cells.

    Science.gov (United States)

    Xu, Linghui; Ding, Wanhong; Stohl, Lori L; Zhou, Xi K; Azizi, Shayan; Chuang, Ethan; Lam, Jimmy; Wagner, John A; Granstein, Richard D

    2017-11-21

    Dermal blood vessels and regional lymph nodes are innervated by sympathetic nerves and, under stress, sympathetic nerves release norepinephrine (NE). Exposure of primary murine dermal microvascular endothelial cells (pDMECs) to NE followed by co-culture with Langerhans cells (LCs), responsive CD4+ T cells and antigen resulted in modulation of CD4+ T cell responses. NE-treatment of pDMECs induced increased production of interleukin (IL)-6 and IL-17A while downregulating interferon (IFN)-γ and IL-22 release. This effect did not require contact between pDMECs and LCs or T cells and depended on pDMEC production of IL-6. The presence of NE-treated pDMECs increased the proportion of CD4+ T cells expressing intracellular IL-17A and increased IL-17A mRNA while decreasing the proportion of IFN-γ- or IL-22-expressing CD4+ T cells and mRNA levels for those cytokines. Retinoic acid receptor-related orphan receptor gamma (RORγt) mRNA was significantly increased in CD4+ T cells while T-box transcription factor (T-bet) mRNA was decreased. Intradermal administration of NE prior to hapten immunization at the injection site produced a similar bias in draining lymph node CD4+ T cells toward IL-17A and away from IFN-γ and IL-22 production. Under stress, release of NE may have significant regulatory effects on the outcome of antigen presentation through actions on ECs as well as enhancement of inflammatory skin disorders involving IL-17/Th17 cells. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  19. Ultrastructure and calcium balance in meristem cells of pea roots exposed to extremely low magnetic fields

    Science.gov (United States)

    Belyavskaya, N. A.

    2001-01-01

    Investigations of low magnetic field (LMF) effects on biological systems have attracted attention of biologists due to planned space flights to other planets where the field intensity does not exceed 10 -5 Oe. Pea ( Pisum sativum L.) seeds were grown in an environment of LMF 3 days. In meristem cells of roots exposed to LMF, one could observe such ultrastructural peculiarities as a noticeable accumulation of lipid bodies, development of a lytic compartment (vacuoles, cytosegresomes and paramural bodies), and reduction of phytoferritin in plastids. Mitochondria were the most sensitive organelle to LMF application. Their size and relative volume in cells increased, matrix was electron-transparent, and cristae reduced. Because of the significant role of calcium signalling in plant responses to different environmental factors, calcium participation in LMF effects was investigated using a pyroantimonate method to identify the localization of free calcium ions. The intensity of cytochemical reaction in root cells after LMF application was strong. The Ca 2+ pyroantimonate deposits were observed both in all organelles and in a hyaloplasm of the cells. Data obtained suggest that the observed LMF effects on ultrastructure of root cells were due to disruptions in different metabolic systems including effects on Ca 2+ homeostasis.

  20. Reduced cytotoxicity in PCB-exposed Chinese Hamster Ovary (CHO) cells pretreated with vitamin E.

    Science.gov (United States)

    Murati, Teuta; Šimić, Branimir; Pleadin, Jelka; Vukmirović, Maja; Miletić, Marina; Durgo, Ksenija; Kniewald, Jasna; Kmetič, Ivana

    2017-01-01

    The aim of this study was to evaluate protective effects of vitamin E (50 -150 μM) in ovary cells upon cytotoxic effects induced by two structurally distinct PCB congeners - planar "dioxin-like" PCB 77 and non-planar di-ortho-substituted PCB 153 with an emphasis on identifying differences in the mechanism of vitamin E action depending on the structure of congeners. Application of three bioassays confirmed that PCBs decrease ovarian cell proliferation with slightly profound effects of PCB 77. PCB - induced ROS production and lipid peroxidation were significant for both congeners with also more noticeable effect for PCB 77. Vitamin E pre-incubation has improved viability of cells, reduced ROS formation and lipid peroxidation induced by PCBs' treatment. Preincubation with vitamin E was more effective when cells where treated with non-planar PCB 153. Altogether, vitamin E action was protective, congener specific and more effective when ovary cells were exposed to ortho-substituted PCB congener. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Pim-3 contributes to radioresistance through regulation of the cell cycle and DNA damage repair in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiang-Yuan; Wang, Zhen [Cancer Research Institute, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Li, Bei [Department of Nuclear Medicine, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Zhang, Ying-Jian, E-mail: yjzhang111@aliyun.com [Department of Nuclear Medicine, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China); Li, Ying-Yi, E-mail: liyingyi@fudan.edu.cn [Cancer Research Institute, Fudan University Shanghai Cancer Center, Department of Oncology, Shanghai Medical College, Fudan University, Shanghai (China)

    2016-04-22

    Resistance of cancer cells to chemoradiotherapy is a major clinical problem in pancreatic cancer treatment. Therefore, understanding the molecular basis of cellular resistance and identifying novel targets are essential for improving treatment efficacy for pancreatic cancer patients. Previous studies have demonstrated a significant role for Pim-3 in pancreatic cancer survival against gemcitabine-induced genotoxic stress. Here, we observed that radiation treatment enhanced Pim-3 expression in human pancreatic cancer cells in vitro. Stable overexpression of Pim-3 in pancreatic cancer cells significantly protected cells against radiation treatment by attenuating G2/M phase cell cycle arrest and DNA damage response. Silencing of Pim-3 expression significantly elevated the phosphorylation of histone variant H2AX, a marker of DNA double strand breaks, and decreased the activation of ataxia-telangiectasia-mutated (ATM) kinase, along with its downstream targets, eventually enhancing the radiosensitivity of human pancreatic cancer cells in vitro and in vivo. Hence, we demonstrated a novel function for Pim-3 in human pancreatic cancer cell survival against radiation. Targeting Pim-3 may be a promising way to improve treatment efficacy in combination with radiotherapy in human pancreatic cancer. - Highlights: • This is first study to demonstrate that Pim-3 is endogenously induced by ionizing radiation in pancreatic cancer cells, and Pim-3 overexpression enhanced radioresistance of pancreatic cancer cells both in vitro and in vivo. • This is first study to provide evidence that radioresistance induced by Pim-3 is mainly attributed to Pim-3 induces activation of ATM, which subsequently activates checkpoint 1, leading to amplification of DNA repair through cell cycle arrest and DNA repair pathways. • This is first study to indicate that targeting Pim-3 may be a promising strategy to provide better treatment efficacy in combination with radiotherapy in human pancreatic

  2. Piperine causes G1 phase cell cycle arrest and apoptosis in melanoma cells through checkpoint kinase-1 activation.

    Directory of Open Access Journals (Sweden)

    Neel M Fofaria

    Full Text Available In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR and checkpoint kinase 1 (Chk1. Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb. Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis.

  3. Piperine causes G1 phase cell cycle arrest and apoptosis in melanoma cells through checkpoint kinase-1 activation.

    Science.gov (United States)

    Fofaria, Neel M; Kim, Sung-Hoon; Srivastava, Sanjay K

    2014-01-01

    In this study, we determined the cytotoxic effects of piperine, a major constituent of black and long pepper in melanoma cells. Piperine treatment inhibited the growth of SK MEL 28 and B16 F0 cells in a dose and time-dependent manner. The growth inhibitory effects of piperine were mediated by cell cycle arrest of both the cell lines in G1 phase. The G1 arrest by piperine correlated with the down-regulation of cyclin D1 and induction of p21. Furthermore, this growth arrest by piperine treatment was associated with DNA damage as indicated by phosphorylation of H2AX at Ser139, activation of ataxia telangiectasia and rad3-related protein (ATR) and checkpoint kinase 1 (Chk1). Pretreatment with AZD 7762, a Chk1 inhibitor not only abrogated the activation of Chk1 but also piperine mediated G1 arrest. Similarly, transfection of cells with Chk1 siRNA completely protected the cells from G1 arrest induced by piperine. Piperine treatment caused down-regulation of E2F1 and phosphorylation of retinoblastoma protein (Rb). Apoptosis induced by piperine was associated with down-regulation of XIAP, Bid (full length) and cleavage of Caspase-3 and PARP. Furthermore, our results showed that piperine treatment generated ROS in melanoma cells. Blocking ROS by tiron protected the cells from piperine mediated cell cycle arrest and apoptosis. These results suggest that piperine mediated ROS played a critical role in inducing DNA damage and activation of Chk1 leading to G1 cell cycle arrest and apoptosis.

  4. Exosomes are released by bystander cells exposed to radiation-induced biophoton signals: Reconciling the mechanisms mediating the bystander effect.

    Directory of Open Access Journals (Sweden)

    Michelle Le

    Full Text Available The objective of our study was to explore a possible molecular mechanism by which ultraviolet (UV biophotons could elicit bystander responses in reporter cells and resolve the problem of seemingly mutually exclusive mechanisms of a physical UV signal & a soluble factor-mediated bystander signal.The human colon carcinoma cell line, HCT116 p53 +/+, was directly irradiated with 0.5 Gy tritium beta particles to induce ultraviolet biophoton emission. Bystander cells were not directly irradiated but were exposed to the emitted UV biophotons. Medium was subsequently harvested from UV-exposed bystander cells. The exosomes extracted from this medium were incubated with reporter cell populations. These reporter cells were then assayed for clonogenic survival and mitochondrial membrane potential with and without prior treatment of the exosomes with RNase.Clonogenic cell survival was significantly reduced in reporter cells incubated with exosomes extracted from cells exposed to secondarily-emitted UV. These exosomes also induced significant mitochondrial membrane depolarization in receiving reporter cells. Conversely, exosomes extracted from non-UV-exposed cells did not produce bystander effects in reporter cells. The treatment of exosomes with RNase prior to their incubation with reporter cells effectively abolished bystander effects in reporter cells and this suggests a role for RNA in mediating the bystander response elicited by UV biophotons and their produced exosomes.This study supports a role for exosomes released from UV biophoton-exposed bystander cells in eliciting bystander responses and also indicates a reconciliation between the UV-mediated bystander effect and the bystander effect which has been suggested in the literature to be mediated by soluble factors.

  5. Exosomes are released by bystander cells exposed to radiation-induced biophoton signals: Reconciling the mechanisms mediating the bystander effect.

    Science.gov (United States)

    Le, Michelle; Fernandez-Palomo, Cristian; McNeill, Fiona E; Seymour, Colin B; Rainbow, Andrew J; Mothersill, Carmel E

    2017-01-01

    The objective of our study was to explore a possible molecular mechanism by which ultraviolet (UV) biophotons could elicit bystander responses in reporter cells and resolve the problem of seemingly mutually exclusive mechanisms of a physical UV signal & a soluble factor-mediated bystander signal. The human colon carcinoma cell line, HCT116 p53 +/+, was directly irradiated with 0.5 Gy tritium beta particles to induce ultraviolet biophoton emission. Bystander cells were not directly irradiated but were exposed to the emitted UV biophotons. Medium was subsequently harvested from UV-exposed bystander cells. The exosomes extracted from this medium were incubated with reporter cell populations. These reporter cells were then assayed for clonogenic survival and mitochondrial membrane potential with and without prior treatment of the exosomes with RNase. Clonogenic cell survival was significantly reduced in reporter cells incubated with exosomes extracted from cells exposed to secondarily-emitted UV. These exosomes also induced significant mitochondrial membrane depolarization in receiving reporter cells. Conversely, exosomes extracted from non-UV-exposed cells did not produce bystander effects in reporter cells. The treatment of exosomes with RNase prior to their incubation with reporter cells effectively abolished bystander effects in reporter cells and this suggests a role for RNA in mediating the bystander response elicited by UV biophotons and their produced exosomes. This study supports a role for exosomes released from UV biophoton-exposed bystander cells in eliciting bystander responses and also indicates a reconciliation between the UV-mediated bystander effect and the bystander effect which has been suggested in the literature to be mediated by soluble factors.

  6. p53 modulates the AMPK inhibitor compound C induced apoptosis in human skin cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Shi-Wei [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Wu, Chun-Ying [Division of Gastroenterology and Hepatology, Taichung Veterans General Hospital, Taichung, Taiwan (China); Wang, Yen-Ting [Department of Medical Research and Education, Cheng Hsin General Hospital, Taipei, Taiwan (China); Kao, Jun-Kai [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Department of Pediatrics, Children' s Hospital, Changhua Christian Hospital, Changhua, Taiwan (China); Lin, Chi-Chen; Chang, Chia-Che; Mu, Szu-Wei; Chen, Yu-Yu [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Chiu, Husan-Wen [Institute of Biotechnology, National Cheng-Kung University, Tainan, Taiwan (China); Agricultural Biotechnology Research Center, Academia Sinica, Taipei, Taiwan (China); Chang, Chuan-Hsun [Department of Surgical Oncology, Cheng Hsin General Hospital, Taipei, Taiwan (China); Department of Nutrition Therapy, Cheng Hsin General Hospital, Taipei, Taiwan (China); School of Nutrition and Health Sciences, Taipei Medical University, Taipei, Taiwan (China); Liang, Shu-Mei [Institute of Biotechnology, National Cheng-Kung University, Tainan, Taiwan (China); Agricultural Biotechnology Research Center, Academia Sinica, Taipei, Taiwan (China); Chen, Yi-Ju [Department of Dermatology, Taichung Veterans General Hospital, Taichung, Taiwan (China); Huang, Jau-Ling [Department of Bioscience Technology, Chang Jung Christian University, Tainan, Taiwan (China); Shieh, Jeng-Jer, E-mail: shiehjj@vghtc.gov.tw [Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Department of Education and Research, Taichung Veterans General Hospital, Taichung, Taiwan (China)

    2013-02-15

    Compound C, a well-known inhibitor of the intracellular energy sensor AMP-activated protein kinase (AMPK), has been reported to cause apoptotic cell death in myeloma, breast cancer cells and glioma cells. In this study, we have demonstrated that compound C not only induced autophagy in all tested skin cancer cell lines but also caused more apoptosis in p53 wildtype skin cancer cells than in p53-mutant skin cancer cells. Compound C can induce upregulation, phosphorylation and nuclear translocalization of the p53 protein and upregulate expression of p53 target genes in wildtype p53-expressing skin basal cell carcinoma (BCC) cells. The changes of p53 status were dependent on DNA damage which was caused by compound C induced reactive oxygen species (ROS) generation and associated with activated ataxia-telangiectasia mutated (ATM) protein. Using the wildtype p53-expressing BCC cells versus stable p53-knockdown BCC sublines, we present evidence that p53-knockdown cancer cells were much less sensitive to compound C treatment with significant G2/M cell cycle arrest and attenuated the compound C-induced apoptosis but not autophagy. The compound C induced G2/M arrest in p53-knockdown BCC cells was associated with the sustained inactive Tyr15 phosphor-Cdc2 expression. Overall, our results established that compound C-induced apoptosis in skin cancer cells was dependent on the cell's p53 status. - Highlights: ► Compound C caused more apoptosis in p53 wildtype than p53-mutant skin cancer cells. ► Compound C can upregulate p53 expression and induce p53 activation. ► Compound C induced p53 effects were dependent on ROS induced DNA damage pathway. ► p53-knockdown attenuated compound C-induced apoptosis but not autophagy. ► Compound C-induced apoptosis in skin cancer cells was dependent on p53 status.

  7. Global gene expression profiling in human lung cells exposed to cobalt

    Energy Technology Data Exchange (ETDEWEB)

    Malard, V.; Berenguer, F.; Prat, O.; Ruat, S.; Steinmetz, G.; Quemeneur, E. [CEA VALRHO, Serv Biochim and Toxicol Nucl, DSV, iBEB, F-30207 Bagnols Sur Ceze (France)

    2007-06-06

    It has been estimated that more than 1 million workers in the United States are exposed to cobalt. Occupational exposure to {sup 59}Co occurs mainly via inhalation and leads to various lung diseases. Cobalt is classified by the IARC as a possible human carcinogen (group 2B). Although there is evidence for in vivo and in vitro toxicity, the mechanisms of cobalt-induced lung toxicity are not fully known. The purpose of this work was to identify potential signatures of acute cobalt exposure using a toxico-genomic approach. Data analysis focused on some cellular processes and protein targets that are thought to be relevant for carcinogenesis, transport and bio-marker research. Results: A time course transcriptome analysis was performed on A549 human pulmonary cells, leading to the identification of 85 genes which are repressed or induced in response to soluble 59 Co. A group of 29 of these genes, representing the main biological functions, was assessed by quantitative RT-PCR. The expression profiles of six of them were then tested by quantitative RT-PCR in a time-dependent manner and three modulations were confirmed by Western blotting. The 85 modulated genes include potential cobalt carriers (FBXL2, ZNT1, SLC12A5), tumor suppressors or transcription factors (MAZ, DLG1, MYC, AXL) and genes linked to the stress response (UBC, HSPCB, BN1P3L). We also identified nine genes coding for secreted proteins as candidates for bio-marker research. Of those, T1MP2 was found to be down-regulated and this modulation was confirmed, in a dose-dependent manner, at protein level in the supernatant of exposed cells. Conclusion: Most of these genes have never been described as related to cobalt stress and provide original hypotheses for further study of the effects of this metal ion on human lung epithelial cells. A putative bio-marker of cobalt toxicity was identified. (authors)

  8. Aneuploidy studies in human cells exposed in vitro to GSM-900 MHz radiofrequency radiation using FISH.

    Science.gov (United States)

    Bourthoumieu, Sylvie; Terro, Faraj; Leveque, Philippe; Collin, Alice; Joubert, Vanessa; Yardin, Catherine

    2011-04-01

    Since previous research found an increase in the rate of aneuploidies in human lymphocytes exposed to radiofrequencies, it seems important to perform further studies. The objective of this study was then to investigate whether the exposure to RF (radiofrequency) radiation similar to that emitted by mobile phones of a second generation standard, i.e., Global System for Mobile communication (GSM) may induce aneuploidy in cultured human cells. The potential induction of genomic instability by GSM-900 MHz radiofrequency (GSM-900) was investigated after in vitro exposure of human amniotic cells for 24 h to average-specific absorption rates (SAR) of 0.25, 1, 2 and 4 W/kg in the temperature range of 36.3-39.7°C. The exposures were carried out in a wire-patch cell (WPC). The rate of aneuploidy of chromosomes 11 and 17 was determined by interphase FISH (Fluorescence In Situ Hybridisation) immediately after independent exposure of three different donors for 24 h. At least 100 interphase cells were analysed per assay. No significant change in the rate of aneuploidy of chromosomes 11 and 17 was found following exposure to GSM-900 for 24 h at average SAR up to 4 W/kg. Our study did not show any in vitro aneuploidogenic effect of GSM using FISH and is not in agreement with the results of previous research.

  9. PPARγ affects nitric oxide in human umbilical vein endothelial cells exposed to Porphyromonas gingivalis.

    Science.gov (United States)

    Li, Peng; Zhang, Dakun; Wan, Meng; Liu, Jianru

    2016-08-01

    Porphyromonas gingivalis induces nitric oxide (NO) synthesis in human umbilical vein endothelial cells (HUVECs). Peroxisome proliferator-activated receptor (PPARγ) has an anti-inflammation function, and its involvement in this NO induction process requires elucidation. Here, we focused on PPARγ expression in HUVECs exposed to P. gingivalis, and investigated its effects on NO synthesis. HUVECs were time-dependently stimulated by P. gingivalis W83 for 0-24h. PPARγ expression was assessed at the mRNA and protein levels, and PPARγ activation was measured using dual-luciferase reporter assays. NO synthesis and NO synthase (NOS) expression in response to P. gingivalis were examined in HUVECs pretreated with representative PPARγ agonist (15-deoxy-Δ12,14-prostaglandin J2 10μM) or antagonist (GW9662 10μM). In addition, NO synthesis and NOS expression in the P. gingivalis infected and control groups were detected. The PPARγ mRNA level in HUVECs increased after exposure to P. gingivalis for 1h and its protein level increased at 2h. Luciferase-induced PPARγ increased in P. gingivalis-exposed HUVECs. NO synthesis in the infected group at 4h, and in the PPARγ-activated group at 8h, was higher than that in controls. Inducible NOS increased in the infected and PPARγ-activated groups at 4 and 8h. The total endothelial NOS (eNOS) and phospho-eNOS levels were lower in the infected group than controls, but did not change in the PPARγ-activated group. Activated PPARγ induces NO generation through the NOS pathway in HUVECs exposed to P. gingivalis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Metallothionein-1 as a biomarker of altered redox metabolism in hepatocellular carcinoma cells exposed to sorafenib.

    Science.gov (United States)

    Houessinon, Aline; François, Catherine; Sauzay, Chloé; Louandre, Christophe; Mongelard, Gaelle; Godin, Corinne; Bodeau, Sandra; Takahashi, Shinichiro; Saidak, Zuzana; Gutierrez, Laurent; Régimbeau, Jean-Marc; Barget, Nathalie; Barbare, Jean-Claude; Ganne, Nathalie; Chauffert, Bruno; Coriat, Romain; Galmiche, Antoine

    2016-05-16

    Sorafenib, a kinase inhibitor active against various solid tumours, induces oxidative stress and ferroptosis, a new form of oxidative necrosis, in some cancer cells. Clinically-applicable biomarkers that reflect the impact of sorafenib on the redox metabolism of cancer cells are lacking. We used gene expression microarrays, real-time PCR, immunoblot, protein-specific ELISA, and gene reporter constructs encoding the enzyme luciferase to study the response of a panel of cancer cells to sorafenib. Tumour explants prepared from surgical hepatocellular carcinoma (HCC) samples and serum samples obtained from HCC patients receiving sorafenib were also used. We observed that genes of the metallothionein-1 (MT1) family are induced in the HCC cell line Huh7 exposed to sorafenib. Sorafenib increased the expression of MT1G mRNA in a panel of human cancer cells, an effect that was not observed with eight other clinically-approved kinase inhibitors. We identified the minimal region of the MT1G promoter that confers inducibility by sorafenib to a 133 base pair region containing an Anti-oxidant Response Element (ARE) and showed the essential role of the transcription factor NRF2 (Nuclear factor erythroid 2-Related Factor 2). We examined the clinical relevance of our findings by analysing the regulation of MT1G in five tumour explants prepared from surgical HCC samples. Finally, we showed that the protein levels of MT1 increase in the serum of some HCC patients receiving sorafenib, and found an association with reduced overall survival. These findings indicate that MT1 constitute a biomarker adapted for exploring the impact of sorafenib on the redox metabolism of cancer cells.

  11. The Bioeffects Resulting from Prokaryotic Cells and Yeast Being Exposed to an 18 GHz Electromagnetic Field.

    Directory of Open Access Journals (Sweden)

    The Hong Phong Nguyen

    Full Text Available The mechanisms by which various biological effects are triggered by exposure to an electromagnetic field are not fully understood and have been the subject of debate. Here, the effects of exposing typical representatives of the major microbial taxa to an 18 GHz microwave electromagnetic field (EMFwere studied. It appeared that the EMF exposure induced cell permeabilisation in all of the bacteria and yeast studied, while the cells remained viable (94% throughout the exposure, independent of the differences in cell membrane fatty acid and phospholipid composition. The resulting cell permeabilisation was confirmed by detection of the uptake of propidium iodine and 23 nm fluorescent silica nanospheres using transmission electron microscopy (TEM and confocal laser scanning microscopy (CLSM. Upon EMF exposure, the bacterial cell membranes are believed to become permeable through quasi-endocytosis processes. The dosimetry analysis revealed that the EMF threshold level required to induce the uptake of the large (46 nm nanopsheres was between three and six EMF doses, with a specific absorption rate (SAR of 3 kW/kg and 5 kW/kg per exposure, respectively, depending on the bacterial taxa being studied. It is suggested that the taxonomic affiliation and lipid composition (e.g. the presence of phosphatidyl-glycerol and/or pentadecanoic fatty acid may affect the extent of uptake of the large nanospheres (46 nm. Multiple 18 GHz EMF exposures over a one-hour period induced periodic anomalous increases in the cell growth behavior of two Staphylococcus aureus strains, namely ATCC 25923 and CIP 65.8T.

  12. The Bioeffects Resulting from Prokaryotic Cells and Yeast Being Exposed to an 18 GHz Electromagnetic Field.

    Science.gov (United States)

    Nguyen, The Hong Phong; Pham, Vy T H; Nguyen, Song Ha; Baulin, Vladimir; Croft, Rodney J; Phillips, Brian; Crawford, Russell J; Ivanova, Elena P

    2016-01-01

    The mechanisms by which various biological effects are triggered by exposure to an electromagnetic field are not fully understood and have been the subject of debate. Here, the effects of exposing typical representatives of the major microbial taxa to an 18 GHz microwave electromagnetic field (EMF)were studied. It appeared that the EMF exposure induced cell permeabilisation in all of the bacteria and yeast studied, while the cells remained viable (94% throughout the exposure), independent of the differences in cell membrane fatty acid and phospholipid composition. The resulting cell permeabilisation was confirmed by detection of the uptake of propidium iodine and 23 nm fluorescent silica nanospheres using transmission electron microscopy (TEM) and confocal laser scanning microscopy (CLSM). Upon EMF exposure, the bacterial cell membranes are believed to become permeable through quasi-endocytosis processes. The dosimetry analysis revealed that the EMF threshold level required to induce the uptake of the large (46 nm) nanopsheres was between three and six EMF doses, with a specific absorption rate (SAR) of 3 kW/kg and 5 kW/kg per exposure, respectively, depending on the bacterial taxa being studied. It is suggested that the taxonomic affiliation and lipid composition (e.g. the presence of phosphatidyl-glycerol and/or pentadecanoic fatty acid) may affect the extent of uptake of the large nanospheres (46 nm). Multiple 18 GHz EMF exposures over a one-hour period induced periodic anomalous increases in the cell growth behavior of two Staphylococcus aureus strains, namely ATCC 25923 and CIP 65.8T.

  13. Transcriptional and secretomic profiling of epidermal cells exposed to alpha particle radiation.

    Science.gov (United States)

    Chauhan, Vinita; Howland, Matthew; Greene, Hillary Boulay; Wilkins, Ruth C

    2012-01-01

    Alpha (α)-particle emitters are probable isotopes to be used in a terrorist attack. The development of biological assessment tools to identify those who have handled these difficult to detect materials would be an asset to our current forensic capacity. In this study, for the purposes of biomarker discovery, human keratinocytes were exposed to α-particle and X-radiation (0.98 Gy/h at 0, 0.5, 1.0, 1.5 Gy) and assessed for differential gene and protein expression using microarray and Bio-Plex technology, respectively. Secretomic analysis of supernatants showed expression of two pro-inflammatory cytokines (IL-13 and PDGF-bb) to be exclusively affected in α-particle exposed cells. The highest dose of α-particle radiation modulated a total of 67 transcripts (fold change>|1.5|, (False discovery rate) FDR2 fold) included KIF20A, NEFM, C7orf10, HIST1H2BD, BMP6, and HIST1H2AC. Among the high expressing genes, five (CCNB2, BUB1, NEK2, CDC20, AURKA) were also differentially expressed at the medium (1.0 Gy) dose however, these genes were unmodulated following exposure to X-irradiation. Networks of these genes clustered around tumor protein-53 and transforming growth factor-beta signaling. This study has identified some potential gene /protein responses and networks that may be validated further to confirm their specificity and potential to be signature biomarkers of α-particle exposure.

  14. Noise Removal with Maintained Spatial Resolution in Raman Images of Cells Exposed to Submicron Polystyrene Particles

    Directory of Open Access Journals (Sweden)

    Linnea Ahlinder

    2016-04-01

    Full Text Available The biodistribution of 300 nm polystyrene particles in A549 lung epithelial cells has been studied with confocal Raman spectroscopy. This is a label-free method in which particles and cells can be imaged without using dyes or fluorescent labels. The main drawback with Raman imaging is the comparatively low spatial resolution, which is aggravated in heterogeneous systems such as biological samples, which in addition often require long measurement times because of their weak Raman signal. Long measurement times may however induce laser-induced damage. In this study we use a super-resolution algorithm with Tikhonov regularization, intended to improve the image quality without demanding an increased number of collected pixels. Images of cells exposed to polystyrene particles have been acquired with two different step lengths, i.e., the distance between pixels, and compared to each other and to corresponding images treated with the super-resolution algorithm. It is shown that the resolution after application of super-resolution algorithms is not significantly improved compared to the theoretical limit for optical microscopy. However, to reduce noise and artefacts in the hyperspectral Raman images while maintaining the spatial resolution, we show that it is advantageous to use short mapping step lengths and super-resolution algorithms with appropriate regularization. The proposed methodology should be generally applicable for Raman imaging of biological samples and other photo-sensitive samples.

  15. Hemorheological characteristics of red blood cells exposed to surface functionalized graphene quantum dots.

    Science.gov (United States)

    Kim, Jeongho; Nafiujjaman, Md; Nurunnabi, Md; Lee, Yong-Kyu; Park, Hun-Kuk

    2016-11-01

    Graphene quantum dots (GQDs) are potential candidates for various biomedical applications such as drug delivery, bioimaging, cell labeling, and biosensors. However, toxicological information on their effects on red blood cells (RBCs) and the mechanisms involved remain unexplored. To the best of our knowledge, our study is the first to investigate the toxicity effects of three GQDs with different surface functionalizations on the hemorheological characteristics of human RBCs, including hemolysis, deformability, aggregation, and morphological changes. RBCs were exposed to three different forms of GQDs (non-functionalized, hydroxylated, and carboxylated GQDs) at various concentrations (0, 500, 750, and 1000 μg/mL) and incubation times (0, 1, 2, 3, or 4 h). The rheological characteristics of the RBCs were measured using microfluidic-laser diffractometry and aggregometry. Overall, the hemolysis rate and rheological alterations of the RBCs were insignificant at a concentration less than 500 μg/mL. Carboxylated GQDs were observed to have more substantial hemolytic activity and caused abrupt changes in the deformability and aggregation of the RBCs than the non-functionalized or hydroxylated GQDs at concentrations >750 μg/mL. Our findings indicate that hemorheological assessments could be utilized to estimate the degree of toxicity to cells and to obtain useful information on safety sheets for nanomaterials. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Oxidative stress response in neural stem cells exposed to different superparamagnetic iron oxide nanoparticles.

    Science.gov (United States)

    Pongrac, Igor M; Pavičić, Ivan; Milić, Mirta; Brkić Ahmed, Lada; Babič, Michal; Horák, Daniel; Vinković Vrček, Ivana; Gajović, Srećko

    2016-01-01

    Biocompatibility, safety, and risk assessments of superparamagnetic iron oxide nanoparticles (SPIONs) are of the highest priority in researching their application in biomedicine. One improvement in the biological properties of SPIONs may be achieved by different functionalization and surface modifications. This study aims to investigate how a different surface functionalization of SPIONs - uncoated, coated with d-mannose, or coated with poly-l-lysine - affects biocompatibility. We sought to investigate murine neural stem cells (NSCs) as important model system for regenerative medicine. To reveal the possible mechanism of toxicity of SPIONs on NSCs, levels of reactive oxygen species, intracellular glutathione, mitochondrial membrane potential, cell-membrane potential, DNA damage, and activities of SOD and GPx were examined. Even in cases where reactive oxygen species levels were significantly lowered in NSCs exposed to SPIONs, we found depleted intracellular glutathione levels, altered activities of SOD and GPx, hyperpolarization of the mitochondrial membrane, dissipated cell-membrane potential, and increased DNA damage, irrespective of the surface coating applied for SPION stabilization. Although surface coating should prevent the toxic effects of SPIONs, our results showed that all of the tested SPION types affected the NSCs similarly, indicating that mitochondrial homeostasis is their major cellular target. Despite the claimed biomedical benefits of SPIONs, the refined determination of their effects on various cellular functions presented in this work highlights the need for further safety evaluations. This investigation helps to fill the knowledge gaps on the criteria that should be considered in evaluating the biocompatibility and safety of novel nanoparticles.

  17. Sildenafil Prevents Apoptosis of Human First-Trimester Trophoblast Cells Exposed to Oxidative Stress

    Science.gov (United States)

    Bolnick, Jay M.; Kilburn, Brian A.; Bolnick, Alan D.; Diamond, Michael P.; Singh, Manvinder; Hertz, Michael; Dai, Jing

    2015-01-01

    Human first-trimester trophoblast cells proliferate at low O2, but survival is compromised by oxidative stress, leading to uteroplacental insufficiency. The vasoactive drug, sildenafil citrate (Viagra, Sigma, St Louis, Missouri), has proven useful in reducing adverse pregnancy outcomes. An important biological function of this pharmaceutical is its action as an inhibitor of cyclic guanosine monophosphate (cGMP) phosphodiesterase type 5 activity, which suggests that it could have beneficial effects on trophoblast survival. To investigate whether sildenafil can prevent trophoblast cell death, human first-trimester villous explants and the HTR-8/SVneo cytotrophoblast cell line were exposed to hypoxia and reoxygenation (H/R) to generate oxidative stress, which induces apoptosis. Apoptosis was optimally inhibited during H/R by 350 ng/mL sildenafil. Sildenafil-mediated survival was reversed by l-NG-nitro-l-arginine methyl ester hydrochloride or cGMP antagonist, indicating a dependence on both nitric oxide (NO) and cGMP. Indeed, either a cGMP agonist or an NO generator was cytoprotective independent of sildenafil. These findings suggest a novel intervention route for patients with recurrent pregnancy loss or obstetrical placental disorders. PMID:25431453

  18. Proteomic profiling of the surface-exposed cell envelope proteins of Caulobacter crescentus.

    Science.gov (United States)

    Cao, Yuan; Bazemore-Walker, Carthene R

    2014-01-31

    Biotinylation of intact cells, avidin enrichment of derivatized peptides, and shotgun proteomics were employed to reveal the composition of the surface-exposed proteome of the aquatic bacterium, Caulobacter crescentus. Ninety-one unique proteins were identified with the majority originating from the outer membrane, periplasm, and inner membrane, subcellular regions that comprise the Gram-negative bacterium cell envelope. Many of these proteins were described as 'conserved hypothetical protein' or 'hypothetical protein'; and so, the actual expression of these gene products was confirmed. Others did not have any known function or lacked annotation. However, this investigation of the Caulobacter surfaceome did reveal the unanticipated presence of a number of enzymes involved in protein degradation. The results presented here can provide a starting point for hypothesis-driven research projects focused on this bacterium in particular and centered on understanding Gram-negative cell architecture and outer membrane biogenesis broadly. The detected protein degradation enzymes anchored on or located within the outer membrane suggest that Caulobacter has nutrient sources larger than small molecules and/or further processes surface proteins once secreted to this location. Additionally, confirmation of outer membrane residency of those proteins predicted to be periplasmic or whose location prediction was not definitive could potentially elucidate the identities of Gram-negative specific anchorless surface proteins. This article is part of a Special Issue entitled: Trends in Microbial Proteomics. © 2013.

  19. Anti-inflammatory activity of the basolateral fraction of Caco-2 cells exposed to a rosemary supercritical extract

    NARCIS (Netherlands)

    Arranz, E.; Mes, J.J.; Wichers, H.J.; Jaime, L.; Reglero, G.; Santoyo, S.

    2015-01-01

    The anti-inflammatory activity of the basolateral fraction of Caco-2 cells exposed to a rosemary supercritical extract was examined. Uptake of rosemary extract fractions was tested on Caco-2 cell monolayers (2–12 h incubation times) and the quantification of carnosic acid and carnosol was performed

  20. ROS-activated ATM-dependent phosphorylation of cytoplasmic substrates identified by large scale phosphoproteomics screen

    DEFF Research Database (Denmark)

    Kozlov, Sergei V; Waardenberg, Ashley J; Engholm-Keller, Kasper

    2016-01-01

    ATM (ataxia-telangiectasia, mutated) protein plays a central role in phosphorylating a network of proteins in response to DNA damage. These proteins function in signalling pathways designed to maintain the stability of the genome and minimize the risk of disease by controlling cell cycle checkpoi......ATM (ataxia-telangiectasia, mutated) protein plays a central role in phosphorylating a network of proteins in response to DNA damage. These proteins function in signalling pathways designed to maintain the stability of the genome and minimize the risk of disease by controlling cell cycle...... to identify cytoplasmic proteins altered in their phosphorylation state in control and A-T (ataxia-telangiectasia) cells in response to oxidative damage. We demonstrated that ATM was activated by oxidative damage in the cytoplasm as well as in the nucleus and identified a total of 9,833 phosphorylation sites...

  1. Alterations in body weight and blood glucose level of female hamsters exposed to electromagnetic fields of cell phones

    Directory of Open Access Journals (Sweden)

    A.R Lotfi

    2010-02-01

    Group 2 was exposed to electromagnetic field emitted by cell phones for 10 days (short term and group 3 for 50 day (long term. In the latter groups, the exposure was 1 hour per day. At the end of the experimental period, the animals were weighed and blood glucose concentrations were determined by obtaining blood samples from 8 randomly selected hamsters in each group.  The blood glucose level was significantly higher in long-term exposed group in comparison with the control and short-term exposed groups (175, 11.6 and 107 mg/dl, respectively (p

  2. Human Invariant Natural Killer T Cells Respond to Antigen-Presenting Cells Exposed to Lipids from Olea europaea Pollen.

    Science.gov (United States)

    Abos Gracia, Beatriz; López Relaño, Juan; Revilla, Ana; Castro, Lourdes; Villalba, Mayte; Martín Adrados, Beatriz; Regueiro, Jose Ramon; Fernández-Malavé, Edgar; Martínez Naves, Eduardo; Gómez Del Moral, Manuel

    2017-01-01

    Allergic sensitization might be influenced by the lipids present in allergens, which can be recognized by natural killer T (NKT) cells on antigen-presenting cells (APCs). The aim of this study was to analyze the effect of olive pollen lipids in human APCs, including monocytes as well as monocyte-derived macrophages (Mϕ) and dendritic cells (DCs). Lipids were extracted from olive (Olea europaea) pollen grains. Invariant (i)NKT cells, monocytes, Mϕ, and DCs were obtained from buffy coats of healthy blood donors, and their cell phenotype was determined by flow cytometry. iNKT cytotoxicity was measured using a lactate dehydrogenase assay. Gene expression of CD1A and CD1D was performed by RT-PCR, and the production of IL-6, IL-10, IL-12, and TNF-α cytokines by monocytes, Mϕ, and DCs was measured by ELISA. Our results showed that monocytes and monocyte-derived Mϕ treated with olive pollen lipids strongly activate iNKT cells. We observed several phenotypic modifications in the APCs upon exposure to pollen-derived lipids. Both Mϕ and monocytes treated with olive pollen lipids showed an increase in CD1D gene expression, whereas upregulation of cell surface CD1d protein occurred only in Mϕ. Furthermore, DCs differentiated in the presence of human serum enhance their surface CD1d expression when exposed to olive pollen lipids. Finally, olive pollen lipids were able to stimulate the production of IL-6 but downregulated the production of lipopolysaccharide- induced IL-10 by Mϕ. Olive pollen lipids alter the phenotype of monocytes, Mϕ, and DCs, resulting in the activation of NKT cells, which have the potential to influence allergic immune responses. © 2017 S. Karger AG, Basel.

  3. Cytogenomics of hexavalent chromium (Cr6+ exposed cells: A comprehensive review

    Directory of Open Access Journals (Sweden)

    Akanksha Nigam

    2014-01-01

    Full Text Available The altered cellular gene expression profile is being hypothesized as the possible molecular basis navigating the onset or progress of various morbidities. This hypothesis has been evaluated here in respect of Cr 6+ induced toxicity. Several studies using gene microarray show selective and strategic dysregulations of cellular genes and pathways induced by Cr 6+ . Relevant literature has been reviewed to unravel these changes in different test systems after exposure to Cr 6+ and also to elucidate association if any, of the altered cytogenomics with Cr 6+ induced toxicity or carcinogenicity. The aim was to verify the hypothesis for critical role of altered cytogenomics in onset of Cr 6+ induced biological / clinical effects by identifying genes modulated commonly by the toxicant irrespective of test system or test concentrations / doses, and by scrutinizing their importance in regulation of the flow of mechanistically linked events crucial for resultant morbidities. Their probability as biomarkers to monitor the toxicant induced biological changes is speculative. The modulated genes have been found to cluster under the pathways that manage onset of oxidative stress, DNA damage, apoptosis, cell-cycle regulation, cytoskeleton, morphological changes, energy metabolism, biosynthesis, oncogenes, bioenergetics, and immune system critical for toxicity. In these studies, the identity of genes has been found to differ remarkably; albeit the trend of pathways′ dysregulation has been found to remain similar. We conclude that the intensity of dysregulation of genes or pathways involved in mechanistic events forms a sub-threshold or threshold level depending upon the dose and type (including speciation of the toxicant, duration of exposure, type of target cells, and niche microenvironment of cells, and the intensity of sub-threshold or threshold level of the altered cytogenomics paves way in toxicant exposed cells eventually either to opt for reversal to

  4. Interphase Death of Chinese Hamster Ovary Cells Exposed to Accelerated Heavy Ions

    Directory of Open Access Journals (Sweden)

    P. Mehnati

    2007-06-01

    Full Text Available Introduction: Heavy ions are nucleus of elements of iron, argon, carbon and neon that all carry positive electrical charges. For these particles to be useful in radiotherapy they need to accelerated to high energy by more than thousand mega volts. Also the cosmic environment is considered to be a complicated mixture of highly energetic photons and heavy ions such as iron. Therefore, the health risks to astronauts during long mission should be considered.  Materials and Methods: The induction of interphase death was tested on Chinese hamster ovary cells by exposing them to accelerated heavy ions (carbon, neon, argon and iron of 10-2000 linear energy transfers (LETs. The fraction of cells that underwent interphase death was determined by observing individual cells with time-lapse photography (direct method as well as by the indirect method of counting cells undergoing interphase death made visible by the addition of caffeine (indirect method. Results: The interphase death due to the exposure to X- rays is increased linearly as the dose exceeds the threshold dose of 10 Gy. Whereas the interphase death increases at a higher rate due to the exposure to high LET heavy ions and no threshold dose was observed. The range of LET values corresponding to the maximum RBE for the interphase death is 120-230 keV/µm. The probability of inducing the interphase death by a single heavy ion traversing through the nucleus is about 0.04-0.08. Discussion and Conclusion: The relative biological effectiveness (RBE of heavy ions as compared to X- rays as determined at the 50% level of induction is increased with LET. It reached a maximum value at a LET of approximately 230 keV/µm and then decreased with further increase in LET. The range of LET values corresponding to the maximum RBE appears to be narrower for interphase death than for reproductive death.

  5. Theoretical analysis of transmembrane potential of cells exposed to nanosecond pulsed electric field.

    Science.gov (United States)

    Lu, Wei; Wu, Ke; Hu, Xiangjun; Xie, Xiangdong; Ning, Jing; Wang, Changzhen; Zhou, Hongmei; Yang, Guoshan

    2017-02-01

    Intracellular electroporation occurs when the cells are exposed to nanosecond pulsed electric field (nsPEF). It is believed the electroporation (formation and extension of pores on the membrane induced by external electric field) is affected significantly by the transmembrane potential. This paper analyzed transmembrane potential induced by nsPEF in the term of pulse frequency spectrum, aiming to provide a theoretical explanation to intracellular bio-effects. Based on the double-shelled spherical cell model, the frequency dependence of transmembrane potential was obtained by solving Laplace's equation, while the time course of transmembrane potential was obtained by a method combined with discrete Fourier transform and Laplace transform. First-order Debye equation was used to describe the dielectric relaxation of the cell medium. Frequency-domain analysis showed that when the electric field frequency was higher than 10 5 Hz, the transmembrane potential on the organelle membrane (ΔΦ o ) was increasing to exceed the transmembrane potential on the cellular membrane (ΔΦ c ). In the time-domain analysis, transmembrane potentials induced by four nsPEF (short trapezoid, long trapezoid, bipolar and sine shapes) with the same field strength were compared with each other. It showed that ΔΦ o is obviously larger than ΔΦ c if the curve of the normalized frequency spectrum of the pulse is more similar with the curve of normalized ΔΦ o in frequency domain. Pulses with major frequency components higher than 10 8 Hz lead to both small ΔΦ o and ΔΦ c . This may explain why high power pulsed microwave lead to unobvious bio-effects of cells than nsPEF with trapezoid form. Through the pulse frequency spectrum it is clearer to understand the relationship between nsPEF and the transmembrane potential.

  6. The protective effect of curcumin in Olfactory Ensheathing Cells exposed to hypoxia.

    Science.gov (United States)

    Bonfanti, Roberta; Musumeci, Teresa; Russo, Cristina; Pellitteri, Rosalia

    2017-02-05

    Curcumin, a phytochemical component derived from the rhizomes of Curcuma longa, has shown a great variety of pharmacological activities, such as anti-inflammatory, anti-tumor, anti-depression and anti-oxidant activity. Therefore, in the last years it has been used as a therapeutic agent since it confers protection in different neurodegenerative diseases, cerebral ischemia and excitotoxicity. Olfactory Ensheathing Cells (OECs) are glial cells of the olfactory system. They are able to secrete several neurotrophic growth factors, promote axonal growth and support the remyelination of damaged axons. OEC transplantation has emerged as a possible experimental therapy to induce repair of spinal cord injury, even if the functional recovery is still limited. Since hypoxia is a secondary effect in spinal cord injury, this in vitro study investigates the protective effect of curcumin in OECs exposed to hypoxia. Primary OECs were obtained from neonatal rat olfactory bulbs and placed both in normal and hypoxic conditions. Furthermore, some cells were grown with basic Fibroblast Growth Factor (bFGF) and/or curcumin at different concentration and times. The results obtained through immunocytochemical procedures and MTT test show that curcumin stimulates cell viability in OECs grown in normal and hypoxic conditions. Furthermore, the synergistic effect of curcumin and bFGF is the most effective exerting protection on OECs. Since spinal cord injury is often accompanied by secondary insults, such as ischemia or hypoxia, our results suggest that curcumin in combination with bFGF might be considered a possible approach for restoration in injuries. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. AT cells are not radiosensitive for simple chromosomal exchanges at low dose

    Energy Technology Data Exchange (ETDEWEB)

    Hada, Megumi; Huff, Janice L.; Patel, Zarana S. [USRA Division of Life Sciences, Houston, TX 77058 (United States); Kawata, Tetsuya [Department of Radiology, School of Medicine, Keio University, Tokyo (Japan); Pluth, Janice M. [Lawrence Berkeley National Laboratory, Life Sciences Division, One Cyclotron Road, Building 74, Berkeley, CA 94720 (United States); George, Kerry A. [Wyle, 1290 Hercules Drive, Houston, TX 77058 (United States); Cucinotta, Francis A., E-mail: Francis.A.Cucinotta@nasa.gov [NASA, Lyndon B. Johnson Space Center, 2101 NASA Parkway, Houston, TX 77058 (United States)

    2011-11-01

    Cells deficient in ATM (product of the gene that is mutated in ataxia telangiectasia patients) or NBS (product of the gene mutated in the Nijmegen breakage syndrome) show increased yields of both simple and complex chromosomal aberrations after high doses (>0.5 Gy) of ionizing radiation (X-rays or {gamma}-rays), however less is known on how these cells respond at low dose. Previously we had shown that the increased chromosome aberrations in ATM and NBS defective lines was due to a significantly larger quadratic dose-response term compared to normal fibroblasts for both simple and complex exchanges. The linear dose-response term for simple exchanges was significantly higher in NBS cells compared to wild type cells, but not for AT cells. However, AT cells have a high background level of exchanges compared to wild type or NBS cells that confounds the understanding of low dose responses. To understand the sensitivity differences for high to low doses, chromosomal aberration analysis was first performed at low dose-rates (0.5 Gy/d), and results provided further evidence for the lack of sensitivity for exchanges in AT cells below doses of 1 Gy. Normal lung fibroblast cells treated with KU-55933, a specific ATM kinase inhibitor, showed increased numbers of exchanges at a dose of 1 Gy and higher, but were similar to wild type cells at 0.5 Gy or below. These results were confirmed using siRNA knockdown of ATM. The present study provides evidence that the increased radiation sensitivity of AT cells for chromosomal exchanges found at high dose does not occur at low dose.

  8. DNA damage in blood cells exposed to low-level lasers.

    Science.gov (United States)

    Sergio, Luiz Philippe da Silva; Silva, Ana Paula Almeida da; Amorim, Philipi Freitas; Campos, Vera Maria Araújo; Magalhães, Luis Alexandre Gonçalves; de Paoli, Flavia; de Souza da Fonseca, Adenilson

    2015-04-01

    In regenerative medicine, there are increasing applications of low-level lasers in therapeutic protocols for treatment of diseases in soft and in bone tissues. However, there are doubts about effects on DNA, and an adequate dosimetry could improve the safety of clinical applications of these lasers. This work aimed to evaluate DNA damage in peripheral blood cells of Wistar rats induced by low-level red and infrared lasers at different fluences, powers, and emission modes according to therapeutic protocols. Peripheral blood samples were exposed to lasers and DNA damage was accessed by comet assay. In other experiments, DNA damage was accessed in blood cells by modified comet assay using formamidopyrimidine DNA glycosylase (Fpg) and endonuclease III enzymes. Data show that exposure to low-level red and infrared lasers induce DNA damage depending on fluence, power and emission mode, which are targeted by Fpg and endonuclease III. Oxidative DNA damage should be considered for therapeutic efficacy and patient safety in clinical applications based on low-level red and infrared lasers. © 2015 Wiley Periodicals, Inc.

  9. Influence of dietary vitamin E on the red cells of ozone-exposed rats

    Energy Technology Data Exchange (ETDEWEB)

    Chow, C.K. (Univ. of Kentucky, Lexington); Kaneko, J.J.

    1979-06-01

    The effect of dietary vitamin E on the susceptibility of red blood cells to ozone exposure was studied in rats. One- and two-month-old male Sprague-Dawley rats were fed a basal vitamin E-deficient diet with or without 45 ppM vitamin E for 4 and 3 months, respectively, and were exposed to 0 or 0.8 ppM ozone continuously for 7 days. Ozone exposure resulted in a significant increase in the activities of glutathione (GSH) peroxidase, pyruvate kinase, and lactate dehydrogenase, and a decrease in GSH level in the red cells of vitamin E-deficient rats, but not in those of the supplemented group. The activities of glucose-6-phosphate dehydrogenase, catalase, and superoxide dismutase and levels of thiobarbituric acid reactants, methemoglobin, hemoglobin, and reticulocytes were not significantly altered by ozone exposure or by the nutritional status of vitamin E. The results suggest that depletion of dietary vitamin E renders animals more susceptible to ozone exposure.

  10. Multiwalled carbon nanotubes enter broccoli cells enhancing growth and water uptake of plants exposed to salinity.

    Science.gov (United States)

    Martínez-Ballesta, M Carmen; Zapata, Lavinia; Chalbi, Najla; Carvajal, Micaela

    2016-06-08

    Carbon nanotubes have been shown to improve the germination and growth of some plant species, extending the applicability of the emerging nano-biotechnology field to crop science. In this work, exploitation of commercial multiwalled carbon nanotubes (MWCNTs) in control and 100 mM NaCl-treated broccoli was performed. Transmission electron microscopy demonstrated that MWCNTs can enter the cells in adult plants with higher accumulation under salt stress. Positive effect of MWCNTs on growth in NaCl-treated plants was consequence of increased water uptake, promoted by more-favourable energetic forces driving this process, and enhanced net assimilation of CO2. MWCNTs induced changes in the lipid composition, rigidity and permeability of the root plasma membranes relative to salt-stressed plants. Also, enhanced aquaporin transduction occurred, which improved water uptake and transport, alleviating the negative effects of salt stress. Our work provides new evidences about the effect of MWCNTs on plasma membrane properties of the plant cell. The positive response to MWCNTs in broccoli plants opens novel perspectives for their technological uses in new agricultural practices, especially when 1plants are exposed to saline environments.

  11. Development of cholesterol granuloma in a temporal bone petrous apex previously containing marrow exposed to air cells.

    Science.gov (United States)

    Selman, Yamil; Wood, John W; Telischi, Fred F; Casiano, Roy R; Angeli, Simon I

    2013-07-01

    There is ongoing debate on the pathogenic mechanisms of cholesterol granuloma formation in the temporal bone. The purpose of this report is to provide evidence in support of the exposed marrow hypothesis in explaining the pathogenesis of petrous apex cholesterol granuloma. Retrospective single case study. The primary outcome evaluated was the diagnosis of a new cholesterol granuloma in a petrous apex that previously demonstrated radiologic evidence of bone marrow exposed to petrous apex air cells. A patient with a unilateral petrous apex cholesterol granuloma develops a new, contralateral cholesterol granuloma in a hyperpneumatized temporal bone petrous apex shown previously to have medullary bone exposed to air cells. This report implicates the medullary-air cell interface in a hyperaerated temporal bone petrous apex in the development and growth of a petrous apex cholesterol granuloma.

  12. The influence of medium conductivity on cells exposed to nsPEF

    Science.gov (United States)

    Moen, Erick K.; Ibey, Bennett L.; Roth, Caleb C.; Barnes, Ronald A.; Beier, Hope T.; Armani, Andrea M.

    2017-02-01

    Nanosecond pulsed electric fields (nsPEF) have proven useful for transporting cargo across cell membranes and selectively activating cellular pathways. The chemistry and biophysics governing this cellular response, however, are complex and not well understood. Recent studies have shown that the conductivity of the solution cells are exposed in could play a significant role in plasma membrane permeabilization and, thus, the overall cellular response. Unfortunately, the means of detecting this membrane perturbation has traditionally been limited to analyzing one possible consequence of the exposure - diffusion of molecules across the membrane. This method has led to contradictory results with respect to the relationship between permeabilization and conductivity. Diffusion experiments also suffer from "saturation conditions" making multi-pulse experiments difficult. As a result, this method has been identified as a key stumbling block to understanding the effects of nsPEF exposure. To overcome these limitations, we recently developed a nonlinear optical imaging technique based on second harmonic generation (SHG) that allows us to identify nanoporation in live cells during the pulse in a wide array of conditions. As a result, we are able to explore and fully test whether lower conductivity extracellular solutions could induce more efficient nanoporation. This hypothesis is based on membrane charging and the relative difference between the extracellular solution and the cytoplasm. The experiments also allow us to test the noise floor of our methodology against the effects of ion leakage. The results emphasize that the electric field, not ionic phenomenon, are the driving force behind nsPEF-induced membrane nanoporation.

  13. Aberrant Gene Expression in Human Non Small Cell Lung Carcinoma Cells Exposed to Demethylating Agent 5-Aza-2'-Deoxycytidine

    Directory of Open Access Journals (Sweden)

    Bao-Zhu Yuan

    2004-07-01

    Full Text Available The identification of genes undergoing genetic or epigenetic alterations and contributing to the development of cancer is critical to our understanding of the molecular mechanisms of carcinogenesis. A new approach in identifying alterations of genes that might be relevant to the process of tumor development was used in this study by examining the gene expression profile in human lung cancer cells exposed to 5-aza-2'deoxycytidine (5-aza-dC. A cDNA array analysis was carried out on 5-aza-dC-treated and untreated non small cell lung cancer (NSCLC cell line NCI-H522. Sixteen and 14 genes were upregulated and downregulated, respectively, by 5-aza-dC treatment. Among them, downregulation of tyrosine protein kinase ABL2 (ABL2 gene and upregulation of hint/protein kinase C inhibitor 1 (Hint/PKCI-1, DVL1, TIMP-1, and TRP-1 genes were found in expanded observations in two or three of five 5-aza-dC-treated NSCLC cell lines. Among these genes, we found that cDNA transfer of Hint/PKCI-1 resulted in a significant in vitro growth inhibition in two cell lines exhibiting 5-aza-dC-induced upregulation of Hint/PKCI-1 and significantly reduced in vivo tumorigenicity of one NSCLC cell line. Hint/PKCI-1, which is the only other characterized human histidine triad (HIT nucleotide-binding protein in addition to tumor-suppressor gene FHIT, might be involved in lung carcinogenesis.

  14. Damage-specific DNA-binding proteins from human cells

    Energy Technology Data Exchange (ETDEWEB)

    Kanjilal, S.

    1992-01-01

    The primary objective of the study was to detect and characterize factors from human cells that bind DNA damaged by ultraviolet radiation. An application of the gel-shift assay was devised in which a DNA probe was UV-irradiated and compared with non-irradiated probe DNA for the ability to bind to such factors in cell extracts. UV-dose dependent binding proteins were identified. Formation of the DNA-protein complexes was independent of the specific sequence, form or source of the DNA. There was a marked preference for lesions on double stranded DNA over those on single stranded DNA. DNA irradiated with gamma rays did not compete with UV-irradiated DNA for the binding activities. Cell lines from patients with genetic diseases associated with disorders of the DNA repair system were screened for the presence of damaged-DNA-binding activities. Simultaneous occurrence of the clinical symptoms of some of these diseases had been previously documented and possible links between the syndromes proposed. However, supporting biochemical or molecular evidence for such associations were lacking. The data from the present investigations indicate that some cases of Xeroderma Pigmentosum group A, Cockayne's Syndrome, Bloom's Syndrome and Ataxia Telangiectasia, all of which exhibit sensitivity to UV or gamma radiation, share an aberrant damaged-DNA-binding factor. These findings support the hypothesis that some of the repair disorder diseases are closely related and may have arisen from a common defect. Partial purification of the binding activities from HeLa cells was achieved. Size-exclusion chromatography resolved the activities into various peaks, one of which was less damage-specific than the others as determined by competition studies using native or UV-irradiated DNA. Some of the activities were further separated by ion-exchange chromatography. On using affinity chromatography methods, the major damage-binding factor could be eluted in the presence of 2 M KCl and 1

  15. Differential responses of neuronal and spermatogenic cells to the doppel cytotoxicity.

    Directory of Open Access Journals (Sweden)

    Kefeng Qin

    Full Text Available Although structurally and biochemically similar to the cellular prion (PrP(C, doppel (Dpl is unique in its biological functions. There are no reports about any neurodegenerative diseases induced by Dpl. However the artificial expression of Dpl in the PrP-deficient mouse brain causes ataxia with Purkinje cell death. Abundant Dpl proteins have been found in testis and depletion of the Dpl gene (Prnd causes male infertility. Therefore, we hypothesize different regulations of Prnd in the nerve and male productive systems. In this study, by electrophoretic mobility shift assays we have determined that two different sets of transcription factors are involved in regulation of the Prnd promoter in mouse neuronal N2a and GC-1 spermatogenic (spg cells, i.e., upstream stimulatory factors (USF in both cells, Brn-3 and Sp1 in GC-1 spg cells, and Sp3 in N2a cells, leading to the expression of Dpl in GC-1 spg but not in N2a cells. We have further defined that, in N2a cells, Dpl induces oxidative stress and apoptosis, which stimulate ataxia-telangiectasia mutated (ATM-modulating bindings of transcription factors, p53 and p21, to Prnp promoter, resulting the PrP(C elevation for counteraction of the Dpl cytotoxicity; in contrast, in GC-1 spg cells, phosphorylation of p21 and N-terminal truncated PrP may play roles in the control of Dpl-induced apoptosis, which may benefit the physiological function of Dpl in the male reproduction system.

  16. Differential Responses of Neuronal and Spermatogenic Cells to the Doppel Cytotoxicity

    Science.gov (United States)

    Qin, Kefeng; Ding, Tianbing; Xiao, Yi; Ma, Wenyu; Wang, Zhen; Gao, Jimin; Zhao, Lili

    2013-01-01

    Although structurally and biochemically similar to the cellular prion (PrPC), doppel (Dpl) is unique in its biological functions. There are no reports about any neurodegenerative diseases induced by Dpl. However the artificial expression of Dpl in the PrP-deficient mouse brain causes ataxia with Purkinje cell death. Abundant Dpl proteins have been found in testis and depletion of the Dpl gene (Prnd) causes male infertility. Therefore, we hypothesize different regulations of Prnd in the nerve and male productive systems. In this study, by electrophoretic mobility shift assays we have determined that two different sets of transcription factors are involved in regulation of the Prnd promoter in mouse neuronal N2a and GC-1 spermatogenic (spg) cells, i.e., upstream stimulatory factors (USF) in both cells, Brn-3 and Sp1 in GC-1 spg cells, and Sp3 in N2a cells, leading to the expression of Dpl in GC-1 spg but not in N2a cells. We have further defined that, in N2a cells, Dpl induces oxidative stress and apoptosis, which stimulate ataxia-telangiectasia mutated (ATM)-modulating bindings of transcription factors, p53 and p21, to Prnp promoter, resulting the PrPC elevation for counteraction of the Dpl cytotoxicity; in contrast, in GC-1 spg cells, phosphorylation of p21 and N-terminal truncated PrP may play roles in the control of Dpl-induced apoptosis, which may benefit the physiological function of Dpl in the male reproduction system. PMID:24339999

  17. Apoptosis of peripheral blood mononuclear cells in children exposed to arsenic and fluoride.

    Science.gov (United States)

    Rocha-Amador, Diana O; Calderón, Jaqueline; Carrizales, Leticia; Costilla-Salazar, Rogelio; Pérez-Maldonado, Iván Nelinho

    2011-11-01

    In this study, we evaluated apoptosis induction in human immune cells in children exposed to arsenic (As) and fluoride (F). Children living in two areas in Mexico (Soledad de Graciano Sanchez (SGS) in San Luis Potosí and Colonia 5 de Febrero in Durango) were studied. Water, urine and blood samples were collected. Approximately 90% of the water samples in 5 de Febrero had As and F levels above the World Health Organization intervention guideline (10 μg/L and 1.5mg/L, respectively). In SGS, 0% of the water samples exceeded Mexican guidelines. Urinary As and F levels in children living in 5 de Febrero were significantly higher than the levels found in children living in SGS. In addition, the level of apoptosis was higher in children from the 5 de Febrero community when compared with the level of apoptosis in children living in SGS. Thus, in a worldwide context, our study demonstrates the health risks to children living in these regions. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. A biosensor of SRC family kinase conformation by exposable tetracysteine useful for cell-based screening.

    Science.gov (United States)

    Irtegun, Sevgi; Wood, Rebecca; Lackovic, Kurt; Schweiggert, Jörg; Ramdzan, Yasmin M; Huang, David C S; Mulhern, Terrence D; Hatters, Danny M

    2014-07-18

    We developed a new approach to distinguish distinct protein conformations in live cells. The method, exposable tetracysteine (XTC), involved placing an engineered tetracysteine motif into a target protein that has conditional access to biarsenical dye binding by conformational state. XTC was used to distinguish open and closed regulatory conformations of Src family kinases. Substituting just four residues with cysteines in the conserved SH2 domain of three Src-family kinases (c-Src, Lck, Lyn) enabled open and closed conformations to be monitored on the basis of binding differences to biarsenical dyes FlAsH or ReAsH. Fusion of the kinases with a fluorescent protein tracked the kinase presence, and the XTC approach enabled simultaneous assessment of regulatory state. The c-Src XTC biosensor was applied in a boutique screen of kinase inhibitors, which revealed six compounds to induce conformational closure. The XTC approach demonstrates new potential for assays targeting conformational changes in key proteins in disease and biology.

  19. Persistent oxidative stress in human neural stem cells exposed to low fluences of charged particles

    Directory of Open Access Journals (Sweden)

    Janet E. Baulch

    2015-08-01

    Full Text Available Exposure to the space radiation environment poses risks for a range of deleterious health effects due to the unique types of radiation encountered. Galactic cosmic rays are comprised of a spectrum of highly energetic nuclei that deposit densely ionizing tracks of damage along the particle trajectory. These tracks are distinct from those generated by the more sparsely ionizing terrestrial radiations, and define the geometric distribution of the complex cellular damage that results when charged particles traverse the tissues of the body. The exquisite radiosensitivity of multipotent neural stem and progenitor cells found within the neurogenic regions of the brain predispose the central nervous system to elevated risks for radiation induced sequelae. Here we show that human neural stem cells (hNSC exposed to different charged particles at space relevant fluences exhibit significant and persistent oxidative stress. Radiation induced oxidative stress was found to be most dependent on total dose rather than on the linear energy transfer of the incident particle. The use of redox sensitive fluorogenic dyes possessing relative specificity for hydroxyl radicals, peroxynitrite, nitric oxide (NO and mitochondrial superoxide confirmed that most irradiation paradigms elevated reactive oxygen and nitrogen species (ROS and RNS, respectively in hNSC over a 1 week interval following exposure. Nitric oxide synthase (NOS was not the major source of elevated nitric oxides, as the use of NOS inhibitors had little effect on NO dependent fluorescence. Our data provide extensive evidence for the capability of low doses of charged particles to elicit marked changes in the metabolic profile of irradiated hNSC. Radiation induced changes in redox state may render the brain more susceptible to the development of neurocognitive deficits that could affect an astronaut’s ability to perform complex tasks during extended missions in deep space.

  20. Attenuation of the DNA Damage Response by Transforming Growth Factor-Beta Inhibitors Enhances Radiation Sensitivity of Non–Small-Cell Lung Cancer Cells In Vitro and In Vivo

    Energy Technology Data Exchange (ETDEWEB)

    Du, Shisuo; Bouquet, Sophie; Lo, Chen-Hao; Pellicciotta, Ilenia; Bolourchi, Shiva [Department of Radiation Oncology, New York University School of Medicine, New York, New York (United States); Parry, Renate [Varian Medical Systems, Palo Alto, California (United States); Barcellos-Hoff, Mary Helen, E-mail: mhbarcellos-hoff@nyumc.org [Department of Radiation Oncology, New York University School of Medicine, New York, New York (United States)

    2015-01-01

    Purpose: To determine whether transforming growth factor (TGF)-β inhibition increases the response to radiation therapy in human and mouse non–small-cell lung carcinoma (NSCLC) cells in vitro and in vivo. Methods and Materials: TGF-β–mediated growth response and pathway activation were examined in human NSCLC NCI-H1299, NCI-H292, and A549 cell lines and murine Lewis lung cancer (LLC) cells. Cells were treated in vitro with LY364947, a small-molecule inhibitor of the TGF-β type 1 receptor kinase, or with the pan-isoform TGF-β neutralizing monoclonal antibody 1D11 before radiation exposure. The DNA damage response was assessed by ataxia telangiectasia mutated (ATM) or Trp53 protein phosphorylation, γH2AX foci formation, or comet assay in irradiated cells. Radiation sensitivity was determined by clonogenic assay. Mice bearing syngeneic subcutaneous LLC tumors were treated with 5 fractions of 6 Gy and/or neutralizing or control antibody. Results: The NCI-H1299, A549, and LLC NSCLC cell lines pretreated with LY364947 before radiation exposure exhibited compromised DNA damage response, indicated by decreased ATM and p53 phosphorylation, reduced γH2AX foci, and increased radiosensitivity. The NCI-H292 cells were unresponsive. Transforming growth factor-β signaling inhibition in irradiated LLC cells resulted in unresolved DNA damage. Subcutaneous LLC tumors in mice treated with TGF-β neutralizing antibody exhibited fewer γH2AX foci after irradiation and significantly greater tumor growth delay in combination with fractionated radiation. Conclusions: Inhibition of TGF-β before radiation attenuated DNA damage recognition and increased radiosensitivity in most NSCLC cells in vitro and promoted radiation-induced tumor control in vivo. These data support the rationale for concurrent TGF-β inhibition and RT to provide therapeutic benefit in NSCLC.

  1. [Instability of chromosomes in human nerve cells (normal and with neuromental diseases)].

    Science.gov (United States)

    Iurov, Iu B; Vorsanova, S G; Solov'ev, I V; Iurov, I Iu

    2010-10-01

    It is assumed that the genetic mechanism of pathogenesis of such widely spread neural and mental diseases as schizophrenia (SZ), autism, ataxia-telangiectasia (AT), and Alzheimer's disease (AD) is associated with structural and functional genomi? instaility in brain cells. Aneuploidy is one of the most important biological markers of genomic instability. The currently available methods of molecular cytogenetics (I-mFISH, QFISH, and ICS-MCB) facilitate the solution of numerous fundamental biological problems, including analysis ofgenomic variations in brain cells. Using these methods, we have studied for the first time aneuploidy in human embryo and adult brain cells (normal and with AT, AD, and SZ) as well as in blood cells of children with autism. The level of aneuploidy was increased two- to threefold in the embryo brain with a subsequent reduction of the number of abnormal cells in the adult brain. In the case of SZ, mosaic aneuploidy for chromosomes 1, 18, and X was found. The study of blood cells from children with autism showed chromosomal mosaicism for chromosomes X, 9, and 15. In the case of AT, we observed a global expression of aneuploidy in up to 20-50% of cortex and cerebellum neurons. In addition, a local instability of chromosome 14 was revealed in the degenerating cerebellum in the form of breaks in the 14q12 region. In the case of AD, a tenfold increase was observed in the level ofaneuploidy for chromosome 21 in brain sections subjected to neurodegeneration. These data indicate that mosaic genomic instability in nerve cells is one of the mechanism of neurodegenerative and mental diseases.

  2. Glycophorin A somatic cell mutation frequencies in Finnish reinforced plastics workers exposed to styrene.

    Science.gov (United States)

    Bigbee, W L; Grant, S G; Langlois, R G; Jensen, R H; Anttila, A; Pfäffli, P; Pekari, K; Norppa, H

    1996-10-01

    We have used the glycophorin A (GPA) in vivo somatic cell mutation assay to assess the genotoxic potential of styrene exposure in 47 reinforced plastics workers occupationally exposed to styrene and 47 unexposed controls matched for age, gender, and active smoking status. GPA variant erythrocyte frequencies (Vf), reflecting GPA allele loss (phi/N) and allele loss and duplication (N/N) somatic mutations arising in vivo in the erythroid progenitor cells of individuals of GPA M/N heterozygous genotype, were flow cytometrically determined in peripheral blood samples from these subjects. Measurements of styrene exposure of the workers at the time of blood sampling showed a mean 8-h time-weighted average (TWA8-h) styrene concentration of 155 mg/m3 (37 ppm) in the breathing zone. Mean urinary concentrations of the styrene metabolites mandelic acid (MA) and mandelic acid plus phenyl glyoxylic acid (MA+PGA) were 4.4 mmol/liter (after workshift) and 2.1 mmol/liter (next morning), respectively. Multivariate analysis of covariance on log-transformed GPA Vf data with models allowing adjustment for age, gender, smoking status, and styrene exposure showed that N/N Vf were nearly significantly increased among all of the exposed workers (adjusted geometric mean, 6.3 per million versus 5.0 in the controls; P = 0.058) and were statistically significantly elevated (adjusted geometric mean, 6.8 versus 5.0 in the controls; P = 0.036) among workers classified into a high-exposure group according to personal TWA8-h concentration of styrene in the breathing zone of > or = 85 mg/m3 (20 ppm; Finnish threshold limit value). Women in this high exposure group showed especially elevated N/N Vf (adjusted geometric mean 8.5 versus 5.3 in control women; P = 0.020); this elevation was also significant if urinary MA+PGA of > or = 1.2 mmol/liter was used as the basis of classification (adjusted geometric mean, 8.3; P = 0.030). The occupational exposure could not be shown to influence phi/N Vf

  3. INCREASED IL-8 AND IL-6 EXPRESSION IN HUMAN AIRWAY EPITHELIAL CELLS EXPOSED TO CARBON ULTRAFINE PARTICLES

    Science.gov (United States)

    INCREASED IL-6 AND IL-8 EXPRESSION IN HUMAN AIRWAY EPITHELIAL CELLS EXPOSED TO CARBON ULTRAFINE PARTICLES.R Silbajoris1, A G Lenz2, I Jaspers3, J M Samet1. 1NHEERL, USEPA, RTP, NC, USA; 2GSF-Institute for Inhalation Biology, Neuherberg, Germany; 3 CEMLB, UNC-CH, Chapel Hill, ...

  4. Unirradiated cells rescue cells exposed to ionizing radiation: Activation of NF-κB pathway in irradiated cells

    Energy Technology Data Exchange (ETDEWEB)

    Lam, R.K.K. [Department of Physics and Materials Science, City University of Hong Kong (Hong Kong); Han, Wei [Center of Medical Physics and Technology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Yu, K.N., E-mail: peter.yu@cityu.edu.hk [Department of Physics and Materials Science, City University of Hong Kong (Hong Kong); State Key Laboratory in Marine Pollution, City University of Hong Kong (Hong Kong)

    2015-12-15

    Highlights: • Rescue effect was observed in both irradiated and HeLa and NIH/3T3 cells. • Novel setup and procedures to separate the rescue signals and the bystander signals. • Confirmed activation of NF-κB pathway in rescue effect using activation inhibitor. • Confirmed activation of NF-κB pathway in rescue effect using anti-NF-κB p65 antibody. - Abstract: We studied the involvement of NF-κB pathway activation in the rescue effect in HeLa and NIH/3T3 cells irradiated by α particles. Firstly, upon irradiation by 5 cGy of α particles, for both cell lines, the numbers of 53BP1 foci/cell at 12 h post-irradiation were significantly smaller when only 2.5% of the cell population was irradiated as compared to 100% irradiation, which demonstrated the rescue effect. Secondly, we studied the effect of NF-κB on the rescue effect through the use of the NF-κB activation inhibitor BAY-11-7082. Novel experimental setup and procedures were designed to prepare the medium (CM) which had conditioned the bystander cells previously partnered with irradiated cells, to ensure physical separation between rescue and bystander signals. BAY-11-7082 itself did not inflict DNA damages in the cells or have effects on activation of the NF-κB response pathway in the irradiated cells through direct irradiation. The rescue effect was induced in both cell lines by the CM, which was abrogated if BAY-11-7082 was added to the CM. Thirdly, we studied the effect of NF-κB on the rescue effect through staining for phosphorylated NF-κB (p-NF-κB) expression using the anti-NF-κB p65 (phospho S536) antibody. When the fraction of irradiated cells dropped from 100% to 2.5%, the p-NF-κB expression in the cell nuclei of irradiated NIH/3T3 cells increased significantly, while that in the cell nuclei of irradiated HeLa cells also increased although not significantly. Moreover, the p-NF-κB expression in the cell nuclei of irradiated HeLa cells and NIH/3T3 cells treated with CM also increased

  5. Increased memory T cell populations in Pb-exposed children from an e-waste-recycling area.

    Science.gov (United States)

    Cao, Junjun; Xu, Xijin; Zhang, Yu; Zeng, Zhijun; Hylkema, Machteld N; Huo, Xia

    2018-03-01

    Chronic exposure to heavy metals could affect cell-mediated immunity. The aim of this study was to explore the status of memory T cell development in preschool children from an e-waste recycling area. Blood lead (Pb) levels, peripheral T cell subpopulations, and serum levels of cytokines (IL-2/IL-7/IL-15), relevant to generation and homeostasis of memory T cells were evaluated in preschool children from Guiyu (e-waste-exposed group) and Haojiang (reference group). The correlations between blood Pb levels and percentages of memory T cell subpopulations were also evaluated. Guiyu children had higher blood Pb levels and increased percentages of CD4 + central memory T cells and CD8 + central memory T cells than in the Haojiang group. Moreover, blood Pb levels were positively associated with the percentages of CD4 + central memory T cells. In contrast, Pb exposure contributed marginally in the change of percentages of CD8 + central memory T cells in children. There was no significant difference in the serum cytokine levels between the e-waste-exposed and reference children. Taken together, preschool children from an e-waste recycling area suffer from relatively higher levels of Pb exposure, which might facilitate the development of CD4 + central memory T cells in these children. Copyright © 2017. Published by Elsevier B.V.

  6. Human chromosome 1 satellite 3 DNA is decondensed, demethylated and transcribed in senescent cells and in A431 epithelial carcinoma cells.

    Science.gov (United States)

    Enukashvily, N I; Donev, R; Waisertreiger, I S-R; Podgornaya, O I

    2007-01-01

    Constitutive heterochromatin mainly consists of different classes of satellite DNAs and is defined as a transcriptionally inactive chromatin that remains compact throughout the cell cycle. The aim of this work was to investigate the level of condensation, methylation and transcriptional status of centromeric (alphoid DNA) and pericentromeric satellites (human satellite 3, HS3) in tissues (lymphocytes, placenta cells) and in cultured primary (MRC5, VH-10, AT2Sp) and malignant (A431) cells. We found that alphoid DNA remained condensed and heavily methylated in all the cell types. The HS3 of chromosome 1 (HS3-1) but not of chromosome 9 (HS3-9) was strongly decondensed and demethylated in A431 cells. The same observation was made for aged embryonic lung (MRC5) and juvenile foreskin (VH-10) fibroblasts obtained at late passages (32(nd) and 23(rd), respectively). Decondensation was also found in ataxia telangiectasia AT2Sp fibroblasts at the 16(th) passage. One of the manifestations of the disease is premature aging. The level of HS3-1 decondensation was higher in aged primary fibroblasts as compared to A431. The HS3-1 extended into the territory of neighbouring chromosomes. An RT-PCR product was detected in A431 and senescent MRC5 fibroblasts using primers specific for HS3-1. The RNA was polyadenylated and transcribed from the reverse chain. Our results demonstrate the involvement of satellite DNA in associations between human chromosomes and intermingling of chromosome territories. The invading satellite DNA can undergo decondensation to a certain level. This process is accompanied by demethylation and transcription. Copyright (c) 2007 S. Karger AG, Basel.

  7. Low-Dose Irradiation Enhances Gene Targeting in Human Pluripotent Stem Cells.

    Science.gov (United States)

    Hatada, Seigo; Subramanian, Aparna; Mandefro, Berhan; Ren, Songyang; Kim, Ho Won; Tang, Jie; Funari, Vincent; Baloh, Robert H; Sareen, Dhruv; Arumugaswami, Vaithilingaraja; Svendsen, Clive N

    2015-09-01

    Human pluripotent stem cells (hPSCs) are now being used for both disease modeling and cell therapy; however, efficient homologous recombination (HR) is often crucial to develop isogenic control or reporter lines. We showed that limited low-dose irradiation (LDI) using either γ-ray or x-ray exposure (0.4 Gy) significantly enhanced HR frequency, possibly through induction of DNA repair/recombination machinery including ataxia-telangiectasia mutated, histone H2A.X and RAD51 proteins. LDI could also increase HR efficiency by more than 30-fold when combined with the targeting tools zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats. Whole-exome sequencing confirmed that the LDI administered to hPSCs did not induce gross genomic alterations or affect cellular viability. Irradiated and targeted lines were karyotypically normal and made all differentiated lineages that continued to express green fluorescent protein targeted at the AAVS1 locus. This simple method allows higher throughput of new, targeted hPSC lines that are crucial to expand the use of disease modeling and to develop novel avenues of cell therapy. The simple and relevant technique described in this report uses a low level of radiation to increase desired gene modifications in human pluripotent stem cells by an order of magnitude. This higher efficiency permits greater throughput with reduced time and cost. The low level of radiation also greatly increased the recombination frequency when combined with developed engineered nucleases. Critically, the radiation did not lead to increases in DNA mutations or to reductions in overall cellular viability. This novel technique enables not only the rapid production of disease models using human stem cells but also the possibility of treating genetically based diseases by correcting patient-derived cells. ©AlphaMed Press.

  8. Evaluation of Trace Elements in Augmentation of Statin-Induced Cytotoxicity in Uremic Serum-Exposed Human Rhabdomyosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Hitoshi Uchiyama

    2018-01-01

    Full Text Available Patients with end-stage kidney disease (ESKD are at higher risk for rhabdomyolysis induced by statin than patients with normal kidney function. Previously, we showed that this increase in the severity of statin-induced rhabdomyolysis was partly due to uremic toxins. However, changes in the quantity of various trace elements in ESKD patients likely contribute as well. The purpose of this study is to determine the effect of trace elements on statin-induced toxicity in rhabdomyosarcoma cells exposed to uremic serum (US cells for a long time. Cell viability, apoptosis, mRNA expression, and intracellular trace elements were assessed by viability assays, flow cytometry, real-time RT-PCR, and ICP-MS, respectively. US cells exhibited greater simvastatin-induced cytotoxicity than cells long-time exposed with normal serum (NS cells (non-overlapping 95% confidence intervals. Intracellular levels of Mg, Mn, Cu, and Zn were significantly less in US cells compared to that in NS cells (p < 0.05 or 0.01. Pre-treatment with TPEN increased simvastatin-induced cytotoxicity and eliminated the distinction between both cells of simvastatin-induced cytotoxicity. These results suggest that Zn deficiencies may be involved in the increased risk for muscle complaints in ESKD patients. In conclusion, the increased severity of statin-induced rhabdomyolysis in ESKD patients may be partly due to trace elements deficiencies.

  9. Exposed hydrophobic residues in human immunodeficiency virus type 1 Vpr helix-1 are important for cell cycle arrest and cell death.

    Directory of Open Access Journals (Sweden)

    R Anthony Barnitz

    Full Text Available The human immunodeficiency virus type 1 (HIV-1 accessory protein viral protein R (Vpr is a major determinant for virus-induced G2/M cell cycle arrest and cytopathicity. Vpr is thought to perform these functions through the interaction with partner proteins. The NMR structure of Vpr revealed solvent exposed hydrophobic amino acids along helices 1 and 3 of Vpr, which could be putative protein binding domains. We previously showed that the hydrophobic patch along helix-3 was important for G2/M blockade and cytopathicity. Mutations of the exposed hydrophobic residues along helix-1 were found to reduce Vpr-induced cell cycle arrest and cell death as well. The levels of toxicity during virion delivery of Vpr correlated with G2/M arrest. Thus, the exposed hydrophobic amino acids in the amino-terminal helix-1 are important for the cell cycle arrest and cytopathicity functions of Vpr.

  10. ATM-deficient human fibroblast cells are resistant to low levels of DNA double-strand break induced apoptosis and subsequently undergo drug-induced premature senescence

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jun; Jo, Yong Hwa; Cho, Chang Hoon; Choe, Wonchae; Kang, Insug; Baik, Hyung Hwan [Department of Biochemistry and Molecular Biology, School of Medicine, Kyung Hee University, 26 Kyunghee-daero, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of); Yoon, Kyung-Sik, E-mail: sky9999@khu.ac.kr [Department of Biochemistry and Molecular Biology, School of Medicine, Kyung Hee University, 26 Kyunghee-daero, Dongdaemun-gu, Seoul 130-701 (Korea, Republic of)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer A-T cells were not hypersensitive to low levels of DNA DSBs. Black-Right-Pointing-Pointer A-T cells have enhanced Akt but defect in activation of p53 and apoptotic proteins. Black-Right-Pointing-Pointer A-T cells underwent premature senescence after DNA damage accumulated. Black-Right-Pointing-Pointer Chemotherapeutic effect in cancer therapy may be associated with premature senescence. -- Abstract: DNA DSBs are induced by IR or radiomimetic drugs such as doxorubicin. It has been indicated that cells from ataxia-telangiectasia patients are highly sensitive to radiation due to defects in DNA repair, but whether they have impairment in apoptosis has not been fully elucidated. A-T cells showed increased sensitivity to high levels of DNA damage, however, they were more resistant to low doses. Normal cells treated with combination of KU55933, a specific ATM kinase inhibitor, and doxorubicin showed increased resistance as they do in a similar manner to A-T cells. A-T cells have higher viability but more DNA breaks, in addition, the activations of p53 and apoptotic proteins (Bax and caspase-3) were deficient, but Akt expression was enhanced. A-T cells subsequently underwent premature senescence after treatment with a low dose of doxorubicin, which was confirmed by G2 accumulation, senescent morphology, and SA-{beta}-gal positive until 15 days repair incubation. Finally, A-T cells are radio-resistant at low doses due to its defectiveness in detecting DNA damage and apoptosis, but the accumulation of DNA damage leads cells to premature senescence.

  11. Chemopreventive properties of pinoresinol-rich olive oil involve a selective activation of the ATM-p53 cascade in colon cancer cell lines.

    Science.gov (United States)

    Fini, Lucia; Hotchkiss, Erin; Fogliano, Vincenzo; Graziani, Giulia; Romano, Marco; De Vol, Edward B; Qin, Huanying; Selgrad, Michael; Boland, C Richard; Ricciardiello, Luigi

    2008-01-01

    The Mediterranean diet is rich in extra virgin olive oil (EVOO) and associated with a lower incidence of colorectal cancer. EVOO contains phenolic extracts with potential anticarcinogenic activity. To assess the anticancer properties of EVOO phenolic extracts using in vitro models. Phenolic profiles of two different EVOOs (A and B) were determined. RKO and HCT116 (both p53 proficient), SW480 (p53 mutant) and HCT116(p53-/-) (p53 knocked out) cell lines were treated with EVOO extracts and assessed for cell viability. Apoptosis was determined by terminal deoxynucleotidyl transferase nick end labeling (TUNEL) assay and changes in Bax transcript levels. Cell cycle analysis was determined by flow cytometry and western blots. To confirm the data, analysis of cell viability and cell cycle was performed with purified pinoresinol. Chemical characterization showed that pinoresinol is the main phenol in EVOO-A, and oleocanthal predominates in EVOO-B. Only EVOO-A affected cell viability, which was significantly more pronounced in p53-proficient cells. At a concentration of 200 nM, p53-proficient cells showed increased apoptosis and G(2)/M arrest. In p53-proficient cells, ataxia telangiectasia mutated (ATM) and its downstream-controlled proteins were upregulated after treatment, with a parallel decrease of cyclin B/cdc2. Identical results on cell viability and cell cycle were obtained with purified pinoresinol, but this required a higher concentration than in EVOO-A. Our results demonstrate that pinoresinol-rich EVOO extracts have potent chemopreventive properties and specifically upregulate the ATM-p53 cascade. This result was achieved at substantially lower concentrations in EVOO than with purified pinoresinol, indicating a possible synergic effect between the various polyphenols in olive oil.

  12. Low CD4+ T-cell levels and B-cell apoptosis in vertically HIV-exposed noninfected children and adolescents.

    Science.gov (United States)

    Miyamoto, Maristela; Pessoa, Silvana D; Ono, Erika; Machado, Daisy M; Salomão, Reinaldo; Succi, Regina C de M; Pahwa, Savita; de Moraes-Pinto, Maria Isabel

    2010-12-01

    Lymphocyte subsets, activation markers and apoptosis were assessed in 20 HIV-exposed noninfected (ENI) children born to HIV-infected women who were or not exposed to antiretroviral (ARV) drugs during pregnancy and early infancy. ENI children and adolescents were aged 6-18 years and they were compared to 25 age-matched healthy non-HIV-exposed children and adolescents (Control). ENI individuals presented lower CD4(+) T cells/mm(3) than Control group (control: 1120.3 vs. ENI: 876.3; t-test, p = 0.030). ENI individuals had higher B-cell apoptosis than Control group (Control: 36.6%, ARV exposed: 82.3%, ARV nonexposed: 68.5%; Kruskal-Wallis, p ENI and in Control children and adolescents. Subtle long-term immune alterations might persist among ENI individuals, but the clinical consequences if any are unknown, and these children require continued monitoring.

  13. Proteomic signature of arabidopsis cell cultures exposed to magnetically induced hyper- and microgravity environments

    NARCIS (Netherlands)

    Herranz, R.; Manzano, A.I.; van Loon, J.J.W.A.; Christianen, P.C.M.; Medina, F.J.

    2013-01-01

    Earth-based microgravity simulation techniques are required due to space research constraints. Using diamagnetic levitation, we exposed Arabidopsis thaliana in vitro callus cultures to environments with different levels of effective gravity and magnetic field strengths (B) simultaneously. The

  14. Disease: H00064 [KEGG MEDICUS

    Lifescience Database Archive (English)

    Full Text Available H00064 Ataxia telangiectasia (AT); Ataxia telangiectasia like disorder (ATLD); Lou...is-Bar syndrome; Boder-Sedgwick syndrome Ataxia-telangiectasia (AT) is an autosomal recessive disorder with

  15. Analyzing gene expression profile in K562 cells exposed to sodium valproate using microarray combined with the connectivity map database.

    Science.gov (United States)

    Zhang, Xiang-Zhong; Yin, Ai-Hua; Lin, Dong-Jun; Zhu, Xiao-Yu; Ding, Qian; Wang, Chun-Huai; Chen, Yun-Xian

    2012-01-01

    To explore the mechanism underlying antileukaemia effect of sodium valproate, the growth and survival of the K562 cell line were investigated. Global profiles of gene expression in K562 cells exposed to sodium valproate were assessed and validated. The differentially expressed genes identified were further used to query the connectivity map database to retrieve a ranked list of compounds that act on the same intracellular targets as sodium valproate. A significant increase in cell apoptosis and a change in gene expression profile were observed in valproate-exposed K562 cells. The significant enrichment analysis of gene ontology terms for the differentially expressed genes showed that these genes were involved in many important biological processes. Eight differentially expressed genes involved in apoptosis were verified by quantitative real-time PCR. The connectivity map analysis showed gene expression profile in K562 cells exposed to sodium valproate was most similar to that of HDACi and PI3K inhibitors, suggesting that sodium valproate might exert antileukaemic action by inhibiting HDAC as well as inhibiting PI3K pathway. In conclusion, our data might provide clues to elucidate the molecular and therapeutic potential of VPA in leukaemia treatment, and the connectivity map is a useful tool for exploring the molecular mechanism of drug action.

  16. Analyzing Gene Expression Profile in K562 Cells Exposed to Sodium Valproate Using Microarray Combined with the Connectivity Map Database

    Directory of Open Access Journals (Sweden)

    Xiang-Zhong Zhang

    2012-01-01

    Full Text Available To explore the mechanism underlying antileukaemia effect of sodium valproate, the growth and survival of the K562 cell line were investigated. Global profiles of gene expression in K562 cells exposed to sodium valproate were assessed and validated. The differentially expressed genes identified were further used to query the connectivity map database to retrieve a ranked list of compounds that act on the same intracellular targets as sodium valproate. A significant increase in cell apoptosis and a change in gene expression profile were observed in valproate-exposed K562 cells. The significant enrichment analysis of gene ontology terms for the differentially expressed genes showed that these genes were involved in many important biological processes. Eight differentially expressed genes involved in apoptosis were verified by quantitative real-time PCR. The connectivity map analysis showed gene expression profile in K562 cells exposed to sodium valproate was most similar to that of HDACi and PI3K inhibitors, suggesting that sodium valproate might exert antileukaemic action by inhibiting HDAC as well as inhibiting PI3K pathway. In conclusion, our data might provide clues to elucidate the molecular and therapeutic potential of VPA in leukaemia treatment, and the connectivity map is a useful tool for exploring the molecular mechanism of drug action.

  17. Dose response of gamma rays and iron nuclei for induction of chromosomal aberrations in normal and repair-deficient cell lines.

    Science.gov (United States)

    George, Kerry A; Hada, Megumi; Jackson, Lori J; Elliott, Todd; Kawata, Tetsuya; Pluth, Janice M; Cucinotta, Francis A

    2009-06-01

    We studied the effects of DNA double-strand break (DSB) repair deficiencies on chromosomal aberration frequency using low doses (gamma rays and high-energy iron ions (LET = 151 keV/microm). Chromosomal aberrations were measured using the fluorescence whole-chromosome painting technique. The cell lines included fibroblasts deficient in ATM (product of the gene that is mutated in ataxia telangiectasia patients) or NBS (product of the gene mutated in the Nijmegen breakage syndrome) and gliomablastoma cells proficient in or lacking DNA-dependent protein kinase (DNA-PK) activity. The yields of both simple and complex chromosomal aberrations were increased in DSB repair-defective cells compared to normal cells; the increase was more than twofold higher for gamma rays compared to iron nuclei. For gamma-ray-induced aberrations, the ATM- and NBS-defective lines were found to have significantly larger quadratic components compared to normal fibroblasts for both simple and complex aberrations, while the linear dose-response term was significantly higher only for the NBS cells. For simple and complex aberrations induced by iron nuclei, regression models preferred purely linear and quadratic dose responses, respectively, for each cell line studied. RBEs were reduced relative to normal cells for all of the DSB repair-defective lines, with the DNA-PK-deficient cells found to have RBEs near unity. The large increase in the quadratic dose-response terms in the DSB repair-deficient cell lines points to the importance of the functions of ATM and NBS in chromatin modifications to facilitate correct DSB repair and to minimize aberration formation. The differences found between AT and NBS cells at lower doses suggest important questions about the applicability of observations of radiation sensitivity at high doses to low-dose exposures.

  18. β-Endorphin Neuronal Cell Transplant Reduces Corticotropin Releasing Hormone Hyperresponse to Lipopolysaccharide and Eliminates Natural Killer Cell Functional Deficiencies in Fetal Alcohol Exposed Rats

    Science.gov (United States)

    Boyadjieva, Nadka I.; Ortigüela, María; Arjona, Alvaro; Cheng, Xiaodong; Sarkar, Dipak K.

    2010-01-01

    Background Natural killer (NK) cell dysfunction is associated with hyperresponse of corticotropin releasing hormone (CRH) to immune challenge and with a loss of β-endorphin (BEP) neurons in fetal alcohol exposed animals. Recently, we established a method to differentiate neural stem cells into BEP neurons using cyclic adenosine monophosphate (cAMP)-elevating agents in cultures. Hence, we determined whether in vitro differentiated BEP neurons could be used for reversing the compromised stress response and immune function in fetal alcohol exposed rats. Methods To determine the effect of BEP neuron transplants on NK cell function, we implanted in vitro differentiated BEP neurons into the paraventricular nucleus of pubertal and adult male rats exposed to ethanol or control in utero. The functionality of transplanted BEP neurons was determined by measuring proopiomelanocortin (POMC) gene expression in these cells and their effects on CRH gene expression under basal and after lipopolysaccaride (LPS) challenge. In addition, the effectiveness of BEP neurons in activating NK cell functions is determined by measuring NK cell cytolytic activity and interferon-γ (IFN-γ) production in the spleen and in the peripheral blood mononuclear cell (PBMC) following cell transplantation. Results We showed here that when these in vitro differentiated BEP neurons were transplanted into the hypothalamus, they maintain biological functions by producing POMC and reducing the CRH neuronal response to the LPS challenge. BEP neuronal transplants significantly increased NK cell cytolytic activity in the spleen and in the PBMC and increased plasma levels of IFN-γ in control and fetal alcohol exposed rats. Conclusions These data further establish the BEP neuronal regulatory role in the control of CRH and NK cell cytolytic function and identify a possible novel therapy to treat stress hyper-response and immune deficiency in fetal alcohol exposed subjects. PMID:19320628

  19. Increase in DNA damage in lymphocytes and micronucleus frequency in buccal cells in silica-exposed workers.

    Science.gov (United States)

    Halder, Ajanta; De, Madhusnata

    2012-01-01

    The alkaline single cell gel electrophoresis (comet assay) was applied to study the genotoxic properties of silica in human peripheral blood lymphocytes (PBL). The study was designed to evaluate the DNA damage of lymphocytes and the end points like micronuclei from buccal smears in a group of 45 workers, occupationally exposed to silica, from small mines and stone quarries. The results were compared to 20 sex and age matched normal individuals. There was a statistically significant difference in the damage levels between the exposed group and the control groups. The types of damages (type I -type 1V) were used to measure the DNA damage. The numbers of micronuclei were higher in the silica-exposed population. The present study suggests that the silica exposure can induce lymphocyte DNA damage and produces significant variation of micronuclei in buccal smear.

  20. Frequency Patterns of T-Cell Exposed Amino Acid Motifs in Immunoglobulin Heavy Chain Peptides Presented by MHCs.

    Science.gov (United States)

    Bremel, Robert D; Homan, E Jane

    2014-01-01

    Immunoglobulins are highly diverse protein sequences that are processed and presented to T-cells by B-cells and other antigen presenting cells. We examined a large dataset of immunoglobulin heavy chain variable regions (IGHV) to assess the diversity of T-cell exposed motifs (TCEMs). TCEM comprise those amino acids in a MHC-bound peptide, which face outwards, surrounded by the MHC histotope, and which engage the T-cell receptor. Within IGHV there is a distinct pattern of predicted MHC class II binding and a very high frequency of re-use of the TCEMs. The re-use frequency indicates that only a limited number of different cognate T-cells are required to engage many different clonal B-cells. The amino acids in each outward-facing TCEM are intercalated with the amino acids of inward-facing MHC groove-exposed motifs (GEM). Different GEM may have differing, allele-specific, MHC binding affinities. The intercalation of TCEM and GEM in a peptide allows for a vast combinatorial repertoire of epitopes, each eliciting a different response. Outcome of T-cell receptor binding is determined by overall signal strength, which is a function of the number of responding T-cells and the duration of engagement. Hence, the frequency of TCEM re-use appears to be an important determinant of whether a T-cell response is stimulatory or suppressive. The frequency distribution of TCEMs implies that somatic hypermutation is followed by T-cell clonal expansion that develops along repeated pathways. The observations of TCEM and GEM derived from immunoglobulins suggest a relatively simple, yet powerful, mechanism to correlate T-cell polyspecificity, through re-use of TCEMs, with a very high degree of specificity achieved by combination with a diversity of GEMs. The frequency profile of TCEMs also points to an economical mechanism for maintaining T-cell memory, recall, and self-discrimination based on an endogenously generated profile of motifs.

  1. Anticlastogenic effect of β-glucan, extracted from Saccharomyces cerevisiae, on cultured cells exposed to ultraviolet radiation

    OpenAIRE

    da Silva, Ariane Fernanda; Oliveira, Rodrigo Juliano; Niwa, Andressa Megumi; D’Epiro, Gláucia Fernanda Rocha; Ribeiro, Lúcia Regina; Mantovani, Mário Sérgio

    2012-01-01

    β-glucan is an important polysaccharide due to its medicinal properties of stimulating the immune system and preventing chronic diseases such as cancer. The aim of the present study was to determine the anticlastogenic effect of β-glucan in cells exposed to ultraviolet radiation (UV). Chromosome aberration assay was performed in drug-metabolizing cells (HTC) and non drug-metabolizing cells (CHO-K1 and repair-deficient CHO-xrs5), using different treatment protocols. Continuous treatment (UV + ...

  2. Human Monocyte-Derived Dendritic Cells Exposed to Microorganisms Involved in Hypersensitivity Pneumonitis Induce a Th1-Polarized Immune Response

    Science.gov (United States)

    Pallandre, Jean-René; Borg, Christophe; Loeffert, Sophie; Gbaguidi-Haore, Houssein; Millon, Laurence

    2013-01-01

    Hypersensitivity pneumonitis (HP) is an immunoallergic disease characterized by a prominent interstitial infiltrate composed predominantly of lymphocytes secreting inflammatory cytokines. Dendritic cells (DCs) are known to play a pivotal role in the lymphocytic response. However, their cross talk with microorganisms that cause HP has yet to be elucidated. This study aimed to investigate the initial interactions between human monocyte-derived DCs (MoDCs) and four microorganisms that are different in nature (Saccharopolyspora rectivirgula [actinomycetes], Mycobacterium immunogenum [mycobacteria], and Wallemia sebi and Eurotium amstelodami [filamentous fungi]) and are involved in HP. Our objectives were to determine the cross talk between MoDCs and HP-causative agents and to determine whether the resulting immune response varied according to the microbial extract tested. The phenotypic activation of MoDCs was measured by the increased expression of costimulatory molecules and levels of cytokines in supernatants. The functional activation of MoDCs was measured by the ability of MoDCs to induce lymphocytic proliferation and differentiation in a mixed lymphocytic reaction (MLR). E. amstelodami-exposed (EA) MoDCs expressed higher percentages of costimulatory molecules than did W. sebi-exposed (WS), S. rectivirgula-exposed (SR), or M. immunogenum-exposed (MI) MoDCs (P < 0.05, Wilcoxon signed-rank test). EA-MoDCs, WS-MoDCs, SR-MoDCs, and MI-MoDCs induced CD4+ T cell proliferation and a Th1-polarized immune response. The present study provides evidence that, although differences were initially observed between MoDCs exposed to filamentous fungi and MoDCs exposed to bacteria, a Th1 response was ultimately promoted by DCs regardless of the microbial extract tested. PMID:23720369

  3. DNA double-strand breaks activate ATM independent of mitochondrial dysfunction in A549 cells.

    Science.gov (United States)

    Kalifa, Lidza; Gewandter, Jennifer S; Staversky, Rhonda J; Sia, Elaine A; Brookes, Paul S; O'Reilly, Michael A

    2014-10-01

    Excessive nuclear or mitochondrial DNA damage can lead to mitochondrial dysfunction, decreased energy production, and increased generation of reactive oxygen species (ROS). Although numerous cell signaling pathways are activated when cells are injured, the ataxia telangiectasia mutant (ATM) protein has emerged as a major regulator of the response to both mitochondrial dysfunction and nuclear DNA double-strand breaks (DSBs). Because mitochondrial dysfunction is often a response to excessive DNA damage, it has been difficult to determine whether nuclear and/or mitochondrial DNA DSBs activate ATM independent of mitochondrial dysfunction. In this study, mitochondrial and nuclear DNA DSBs were generated in the A549 human lung adenocarcinoma cell line by infecting with retroviruses expressing the restriction endonuclease PstI fused to a mitochondrial targeting sequence (MTS) or nuclear localization sequence (NLS) and a hemagglutinin antigen epitope tag (HA). Expression of MTS-PstI-HA or NLS-PstI-HA activated the DNA damage response defined by phosphorylation of ATM, the tumor suppressor protein p53 (TP53), KRAB-associated protein (KAP)-1, and structural maintenance of chromosomes (SMC)-1. Phosphorylated ATM and SMC1 were detected in nuclear fractions, whereas phosphorylated TP53 and KAP1 were detected in both mitochondrial and nuclear fractions. PstI also enhanced expression of the cyclin-dependent kinase inhibitor p21 and inhibited cell growth. This response to DNA damage occurred in the absence of detectable mitochondrial dysfunction and excess production of ROS. These findings reveal that DNA DSBs are sufficient to activate ATM independent of mitochondrial dysfunction and suggest that the activated form of ATM and some of its substrates are restricted to the nuclear compartment, regardless of the site of DNA damage. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Identification of genes differentially expressed between benign and osteopontin transformed rat mammary epithelial cells

    Directory of Open Access Journals (Sweden)

    Rudland Philip S

    2009-02-01

    Full Text Available Abstract Background Osteopontin is a secreted, integrin-binding and phosphorylated acidic glycoprotein which has an important role in tumor progression. Findings In this study, we have utilized suppressive subtractive hybridization (SSH to evaluate OPN regulated gene expression, using the Rama 37 benign non-invasive rat mammary cell line and a subclone, Rama 37-OPN. Rama 37-OPN was produced by stably transfecting Rama 37 with an OPN expression vector and it demonstrates increased malignant properties in vitro. Sequence and expression array analysis of the respective cDNA libraries of over 1600 subtracted cDNA fragments revealed 982 ESTs, 45 novel sequences and 659 known genes. The known up-regulated genes in the Rama 37-OPN library code for proteins with a variety of functions including those involved in metabolism, cell adhesion and migration, signal transduction and in apoptosis. Four of the most differentially expressed genes between the benign and in vitro malignant rat mammary cell lines are tumor protein translationally controlled I (TPTI, aryl hydrocarbon receptor nuclear translocator (ARNT, ataxia telangiectasia mutated (ATM and RAN GTPase (RAN. The largest difference (ca 10,000 fold between the less aggressively (MCF-7, ZR-75 and more aggressively malignant (MDA MB 231, MDA MB 435S human breast cancer cell lines is that due to RAN, the next is that due to osteopontin itself. Conclusion The results suggest that enhanced properties associated with the malignant state in vitro induced by osteopontin may be due to, in part, overexpression of RAN GTPase and these biological results are the subject of a subsequent publication 1.

  5. miR-181a promotes G1/S transition and cell proliferation in pediatric acute myeloid leukemia by targeting ATM.

    Science.gov (United States)

    Liu, Xiaodan; Liao, Wang; Peng, Hongxia; Luo, Xuequn; Luo, Ziyan; Jiang, Hua; Xu, Ling

    2016-01-01

    Abnormal expression of miRNAs is intimately related to a variety of human cancers. The purpose of this study is to confirm the expression of miR-181a and elucidate its physiological function and mechanism in pediatric acute myeloid leukemia (AML). Pediatric AML patients and healthy controls were enrolled, and the expression of miR-181a and ataxia telangiectasia mutated (ATM) in tissues were examined using quantitative PCR. Moreover, cell proliferation and cell cycle were evaluated in several cell lines (HL60, NB4 and K562) by using flow cytometry after transfected with miR-181a mimics and inhibitors, or ATM siRNA and control siRNA. Finally, ATM as the potential target protein of miR-181a was examined. We found that miR-181a was significantly increased in pediatric AML, which showed an inverse association with ATM expression. Overexpressed miR-181a in cell lines significantly enhanced cell proliferation, as well as increased the ratio of S-phase cells by miR-181a mimics transfection in vitro. Luciferase activity of the reporter construct identified ATM as the direct molecular target of miR-181a. ATM siRNA transfection significantly enhanced cell proliferation and increased the ratio of S-phase cells in vitro. The results revealed novel mechanism through which miR-181a regulates G1/S transition and cell proliferation in pediatric AML by regulating the tumor suppressor ATM, providing insights into the molecular mechanism in pediatric AML.

  6. Differential protein expression in two bivalve species; Mytilus galloprovincialis and Corbicula fluminea; exposed to Cylindrospermopsis raciborskii cells.

    Science.gov (United States)

    Puerto, Maria; Campos, Alexandre; Prieto, Ana; Cameán, Ana; de Almeida, André Martinho; Coelho, Ana Varela; Vasconcelos, Vitor

    2011-01-17

    The cyanobacteria Cylindrospermopsis raciborskii is considered a threat to aquatic organisms due to the production of the toxin cylindrospermopsin (CYN). Despite the numerous reports evidencing the toxic effects of C. raciborskii cells and CYN in different species, not much is known regarding the toxicity mechanisms associated with this toxin and the cyanobacteria. In this work, a proteomics approach based in the two-dimensional gel electrophoresis and mass spectrometry was used to study the effects of the exposure of two bivalve species, Mytilus galloprovincialis and Corbicula fluminea, to CYN producing (CYN+) and non-producing (CYN-) C. raciborskii cells. Additionally the activities of glutathione S-transferase (GST) and glutathione peroxidase (GPx) were determined. Alterations in actin and tubulin isoforms were detected in gills of both bivalve species and digestive gland of M. galloprovincialis when exposed to CYN- and CYN+ cells. Moreover, GST and GPx activities changed in gills and digestive tract of bivalves exposed to both C. raciborskii freeze dried cells, in comparison to control animals exposed to the green alga Chlorella vulgaris. These results suggest the induction of physiological stress and tissue injury in bivalves by C. raciborskii. This condition is supported by the changes observed in GPx and GST activities which indicate alterations in the oxidative stress defense mechanisms. The results also evidence the capacity of CYN non-producing C. raciborskii to induce biochemical responses and therefore its toxicity potential to bivalves. The heat shock protein 60 (HSP60), extrapallial (EP) fluid protein and triosephosphate isomerase homologous proteins from gills of M. galloprovincialis were down-regulated specifically with the presence of CYN+ C. raciborskii cells. The presence of CYN may lead to additional toxic effects in M. galloprovincialis. This work demonstrates that proteomics is a powerful approach to characterize the biochemical effects of C

  7. Evaluation of cytotoxicity, morphological alterations and oxidative stress in Chinook salmon cells exposed to copper oxide nanoparticles.

    Science.gov (United States)

    Srikanth, Koigoora; Pereira, Eduarda; Duarte, Armando C; Rao, Janapala Venkateswara

    2016-05-01

    The current study is aimed to study cytotoxicity and oxidative stress mediated changes induced by copper oxide nanoparticles (CuO NPs) in Chinook salmon cells (CHSE-214). To this end, a number of biochemical responses are evaluated in CHSE-214 cells which are as follows [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide] MTT, neutral red uptake (NRU), lactate dehydrogenase (LDH), protein carbonyl (PC), lipid peroxidation (LPO), oxidised glutathione (GSSG), reduced glutathione (GSH), glutathione peroxidase (GPx), glutathione sulfo-transferase (GST), superoxide dismutase (SOD), catalase (CAT), 8-Hydroxy-2'-deoxyguanosine (8-OHdG) and reactive oxygen species (ROS), respectively. The 50% inhibition concentration (IC50) of CuO NPs to CHSE-214 cells after 24 h exposure was found to be 19.026 μg ml(-1). Viability of cells was reduced by CuO NPs, and the decrease was dose dependent as revealed by the MTT and NRU assay. CHSE-214 cells exposed to CuO NPs induced morphological changes. Initially, cells started to detach from the surface (12 h), followed by polyhedric, fusiform appearance (19 h) and finally the cells started to shrink. Later, the cells started losing their cellular contents leading to their death only after 24 h. LDH, PC, LPO, GSH, GPx, GST, SOD, CAT, 8-OHdG and ROS responses were seen significantly increased with the increase in the concentration of CuO NPs when compared to their respective controls. However, significant decrease in GSSG was perceptible in CHSE-214 cells exposed to CuO NPs in a dose-dependent manner. Our data demonstrated that CuO NPs induced cytotoxicity in CHSE-214 cells through the mediation of oxidative stress. The current study provides a baseline for the CuO NPs-mediated cytotoxic assessment in CHSE-214 cells for the future studies.

  8. Phenotypic malignant changes and untargeted lipidomic analysis of long-term exposed prostate cancer cells to endocrine disruptors

    Energy Technology Data Exchange (ETDEWEB)

    Bedia, Carmen, E-mail: carmen.bedia@idaea.csic.es; Dalmau, Núria, E-mail: nuria.dalmau@idaea.csic.es; Jaumot, Joaquim, E-mail: joaquim.jaumot@idaea.csic.es; Tauler, Romà, E-mail: roma.tauler@idaea.csic.es

    2015-07-15

    Endocrine disruptors (EDs) are a class of environmental toxic molecules able to interfere with the normal hormone metabolism. Numerous studies involve EDs exposure to initiation and development of cancers, including prostate cancer. In this work, three different EDs (aldrin, aroclor 1254 and chlorpyrifos (CPF)) were investigated as potential inducers of a malignant phenotype in DU145 prostate cancer cells after a chronic exposure. Epithelial to mesenchymal transition (EMT) induction, proliferation, migration, colony formation and release of metalloproteinase 2 (MMP-2) were analyzed in 50-day exposed cells to the selected EDs. As a result, aldrin and CPF exposure led to an EMT induction (loss of 16% and 14% of E-cadherin levels, respectively, compared to the unexposed cells). Aroclor and CPF presented an increased migration (134% and 126%, respectively), colony formation (204% and 144%, respectively) and MMP-2 release (137% in both cases) compared to the unexposed cells. An untargeted lipidomic analysis was performed to decipher the lipids involved in the observed transformations. As general results, aldrin exposure showed a global decrease in phospholipids and sphingolipids, and aroclor and CPF showed an increase of certain phospholipids, glycosphingolipids as well as a remarkable increase of some cardiolipin species. Furthermore, the three exposures resulted in an increase of some triglyceride species. In conclusion, some significant changes in lipids were identified and thus we postulate that some lipid compounds and lipid metabolic pathways could be involved in the acquisition of the malignant phenotype in exposed prostate cancer cells to the selected EDs. - Highlights: • Aldrin, aroclor and chlorpyrifos induced an aggressive phenotype in DU145 cells. • An untargeted lipidomic analysis has been performed on chronic exposed cells. • Lipidomic results showed changes in specific lipid species under chronic exposure. • These lipids may have a role in the

  9. Sodium pyruvate modulates cell death pathways in HaCaT keratinocytes exposed to half-mustard gas.

    Science.gov (United States)

    Paromov, Victor; Brannon, Marianne; Kumari, Sudha; Samala, Mallikarjun; Qui, Min; Smith, Milton; Stone, William L

    2011-03-01

    2-Chloroethyl ethyl sulfide (CEES) or half-mustard gas, a sulfur mustard (HD) analog, is a genotoxic agent that causes oxidative stress and induces both apoptotic and necrotic cell death. Sodium pyruvate induced a necrosis-to-apoptosis shift in HaCaT cells exposed to CEES levels ≤ 1.5 mmol/L and lowered markers of DNA damage, oxidative stress, and inflammation. This study provides a rationale for the future development of multicomponent therapies for HD toxicity in the skin. We hypothesize that a combination of pyruvates with scavengers/antioxidants encapsulated in liposomes for optimal local delivery should be therapeutically beneficial against HD-induced skin injury. However, the latter suggestion should be verified in animal models exposed to HD.

  10. Virus-specific nucleic acids in SV40-exposed hamster embryo cell lines: correlation with S and T antigens.

    Science.gov (United States)

    Levin, M J; Oxman, M N; Diamandopoulos, G T; Levine, A S; Henry, P H; Enders, J F

    1969-02-01

    A number of homologous SV40-exposed hamster embryonic cell lines were examined for the presence of RNA complementary to SV40 DNA. Only those lines containing the SV40 T antigen were found to have such virus-specific RNA. In lines containing the SV40 S antigen, but not the SV40 T antigen, virus-specific RNA was not detected. These findings suggest that the S antigen is not coded for directly by the SV40 genome.

  11. VIRUS-SPECIFIC NUCLEIC ACIDS IN SV40-EXPOSED HAMSTER EMBRYO CELL LINES: CORRELATION WITH S AND T ANTIGENS*

    Science.gov (United States)

    Levin, Myron J.; Oxman, Michael N.; Diamandopoulos, George Th.; Levine, Arthur S.; Henry, Patrick H.; Enders, John F.

    1969-01-01

    A number of homologous SV40-exposed hamster embryonic cell lines were examined for the presence of RNA complementary to SV40 DNA. Only those lines containing the SV40 T antigen were found to have such virus-specific RNA. In lines containing the SV40 S antigen, but not the SV40 T antigen, virus-specific RNA was not detected. These findings suggest that the S antigen is not coded for directly by the SV40 genome. PMID:4307716

  12. Gene expression profiling and pathway analysis of human bronchial epithelial cells exposed to airborne particulate matter collected from Saudi Arabia

    OpenAIRE

    Sun, Hong; Shamy, Magdy; Kluz, Thomas; Muñoz, Alexandra B.; Zhong, Mianhua; Laulicht, Freda; Alghamdi, Mansour A.; Khoder, Mamdouh I; Chen, Lung-Chi; Costa, Max

    2012-01-01

    Epidemiological studies have established a positive correlation between human mortality and increased concentration of airborne particulate matters (PM). However, the mechanisms underlying PM related human diseases, as well as the molecules and pathways mediating the cellular response to PM, are not fully understood. This study aims to investigate the global gene expression changes in human cells exposed to PM10 and to identify genes and pathways that may contribute to PM related adverse heal...

  13. REV-ERBα inhibits the PTGS2 expression in bovine uterus endometrium stromal and epithelial cells exposed to ovarian steroids.

    Science.gov (United States)

    Isayama, Keishiro; Chen, Huatao; Yamauchi, Nobuhiko; Hattori, Masa-Aki

    2014-01-01

    The nuclear receptor REV-ERBα (encoded by NR1D1) has a critical role in metabolism and physiology as well as circadian rhythm. Here, we investigated the possible contribution of clock genes including NR1D1 to the secretion of prostaglandin F2α (PGF2α) from bovine uterine stromal (USCs) and epithelial cells (UECs) by modulating the expression of PTGS2. The circadian oscillation of clock genes in the cells was weak compared with that reported in rodents, but the expression of BMAL1, PER1, and NR1D1 was changed temporally by treatment with ovarian steroids. Significant expression of clock genes including NR1D1 was detected in USCs exposed to progesterone. NR1D1 was also significantly expressed in UECs exposed to estradiol. The expression of PTGS2 was suppressed in USCs exposed to progesterone, while the expression was initially suppressed in UECs exposed to estradiol and then increased after long-term exposure to estradiol. BMAL1 knockdown with specific siRNA caused a significant decrease in the transcript levels of NR1D1 and PTGS2 in USCs, but not in UECs. The production of PGF2α also decreased in USCs after BMAL1 knockdown, while its level did not significantly change in UECs. The transcript level of PTGS2 was increased by treatment with the antagonist of REV-ERBα in both cell types, but the agonist was ineffective. In these two cell types treated with the agonist or antagonist, the PGF2α production coincided well with the PTGS2 expression. Collectively, these results indicate that REV-ERBα plays an inhibitory role in the expression of PTGS2 in both bovine USCs and UECs treated with ovarian steroids.

  14. Gene expression profiling and pathway analysis of human bronchial epithelial cells exposed to airborne particulate matter collected from Saudi Arabia.

    Science.gov (United States)

    Sun, Hong; Shamy, Magdy; Kluz, Thomas; Muñoz, Alexandra B; Zhong, Mianhua; Laulicht, Freda; Alghamdi, Mansour A; Khoder, Mamdouh I; Chen, Lung-Chi; Costa, Max

    2012-12-01

    Epidemiological studies have established a positive correlation between human mortality and increased concentration of airborne particulate matters (PM). However, the mechanisms underlying PM related human diseases, as well as the molecules and pathways mediating the cellular response to PM, are not fully understood. This study aims to investigate the global gene expression changes in human cells exposed to PM(10) and to identify genes and pathways that may contribute to PM related adverse health effects. Human bronchial epithelial cells were exposed to PM(10) collected from Saudi Arabia for 1 or 4 days, and whole transcript expression was profiled using the GeneChip human gene 1.0 ST array. A total of 140 and 230 genes were identified that significantly changed more than 1.5 fold after PM(10) exposure for 1 or 4 days, respectively. Ingenuity Pathway Analysis revealed that different exposure durations triggered distinct pathways. Genes involved in NRF2-mediated response to oxidative stress were up-regulated after 1 day exposure. In contrast, cells exposed for 4 days exhibited significant changes in genes related to cholesterol and lipid synthesis pathways. These observed changes in cellular oxidative stress and lipid synthesis might contribute to PM related respiratory and cardiovascular disease. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. ATM Expression Predicts Veliparib and Irinotecan Sensitivity in Gastric Cancer by Mediating P53-Independent Regulation of Cell Cycle and Apoptosis.

    Science.gov (United States)

    Subhash, Vinod Vijay; Tan, Shi Hui; Yeo, Mei Shi; Yan, Fui Leng; Peethala, Praveen C; Liem, Natalia; Krishnan, Vaidehi; Yong, Wei Peng

    2016-12-01

    Identification of synthetically lethal cellular targets and synergistic drug combinations is important in cancer chemotherapy as they help to overcome treatment resistance and increase efficacy. The Ataxia Telangiectasia Mutated (ATM) kinase is a nuclear protein that plays a major role in the initiation of DNA repair signaling and cell-cycle check points during DNA damage. Although ATM was shown to be associated with poor prognosis in gastric cancer, its implications as a predictive biomarker for cancer chemotherapy remain unexplored. The present study evaluated ATM-induced synthetic lethality and its role in sensitization of gastric cancer cells to PARP and TOP1 inhibitors, veliparib (ABT-888) and irinotecan (CPT-11), respectively. ATM expression was detected in a panel of gastric cell lines, and the IC50 against each inhibitors was determined. The combinatorial effect of ABT-888 and CPT-11 in gastric cancer cells was also determined both in vitro and in vivo ATM deficiency was found to be associated with enhanced sensitivity to ABT-888 and CPT-11 monotherapy, hence suggesting a mechanism of synthetic lethality. Cells with high ATM expression showed reduced sensitivity to monotherapy; however, they showed a higher therapeutic effect with ABT-888 and CPT-11 combinatorial therapy. Furthermore, ATM expression was shown to play a major role in cellular homeostasis by regulating cell-cycle progression and apoptosis in a P53-independent manner. The present study highlights the clinical utility of ATM expression as a predictive marker for sensitivity of gastric cancer cells to PARP and TOP1 inhibition and provides a deeper mechanistic insight into ATM-dependent regulation of cellular processes. Mol Cancer Ther; 15(12); 3087-96. ©2016 AACR. ©2016 American Association for Cancer Research.

  16. Impact of lysosomal storage disorders on biology of mesenchymal stem cells: Evidences from in vitro silencing of glucocerebrosidase (GBA) and alpha-galactosidase A (GLA) enzymes.

    Science.gov (United States)

    Squillaro, Tiziana; Antonucci, Ivana; Alessio, Nicola; Esposito, Anna; Cipollaro, Marilena; Melone, Mariarosa Anna Beatrice; Peluso, Gianfranco; Stuppia, Liborio; Galderisi, Umberto

    2017-12-01

    Lysosomal storage disorders (LDS) comprise a group of rare multisystemic diseases resulting from inherited gene mutations that impair lysosomal homeostasis. The most common LSDs, Gaucher disease (GD), and Fabry disease (FD) are caused by deficiencies in the lysosomal glucocerebrosidase (GBA) and alpha-galactosidase A (GLA) enzymes, respectively. Given the systemic nature of enzyme deficiency, we hypothesized that the stem cell compartment of GD and FD patients might be also affected. Among stem cells, mesenchymal stem cells (MSCs) are a commonly investigated population given their role in hematopoiesis and the homeostatic maintenance of many organs and tissues. Since the impairment of MSC functions could pose profound consequences on body physiology, we evaluated whether GBA and GLA silencing could affect the biology of MSCs isolated from bone marrow and amniotic fluid. Those cell populations were chosen given the former's key role in organ physiology and the latter's intriguing potential as an alternative stem cell model for human genetic disease. Our results revealed that GBA and GLA deficiencies prompted cell cycle arrest along with the impairment of autophagic flux and an increase of apoptotic and senescent cell percentages. Moreover, an increase in ataxia-telangiectasia-mutated staining 1 hr after oxidative stress induction and a return to basal level at 48 hr, along with persistent gamma-H2AX staining, indicated that MSCs properly activated DNA repair signaling, though some damages remained unrepaired. Our data therefore suggest that MSCs with reduced GBA or GLA activity are prone to apoptosis and senescence due to impaired autophagy and DNA repair capacity. © 2017 Wiley Periodicals, Inc.

  17. Signs of Müller cell gliotic response found in the retina of newts exposed to real and simulated microgravity

    Science.gov (United States)

    Grigoryan, E. N.; Anton, H. J.; Poplinskaya, V. A.; Aleinikova, K. S.; Domaratskaya, E. I.; Novikova, Y. P.; Almeida, E.

    2012-05-01

    The effects of real and simulated microgravity on the eye tissue regeneration of newts were investigated. For the first time changes in Müller glial cells in the retina of eyes regenerating after retinal detachment were detected in newts exposed to clinorotation. The cells divided, were hypertrophied, and their processes were thickened. Such changes suggested reactive gliosis and were more significant in animals exposed to rotation when compared with desk-top controls. Later experiments onboard the Russian biosatellite Bion-11 showed similar changes in the retinas that were regenerating in a two-week spaceflight. In the Bion-11 animals, GFAP, the major structural protein of retinal macroglial cells, was found to be upregulated. In a more recent experiment onboard Foton-M3 (2007), GFAP expression in retinas of space-flown, ground control (kept at 1 g), and basal control (sacrificed on launch day) newts was quantified, using microscopy, immunohistochemistry, and digital image analysis. A low level of immunoreactivity was observed in basal controls. In contrast, retinas of space-flown animals showed greater GFAP immunoreactivity associated with both an increased cell number and a higher thickness of intermediate filaments. This, in turn, was accompanied by up-regulation of stress protein (HSP90) and growth factor (FGF2) expressions. It can be postulated that such a response of Müller cells was to mitigate the retinal stress in newts exposed to microgravity. Taken together, the data suggest that the retinal population of macroglial cells could be sensitive to gravity changes and that in space it can react by enhancing its neuroprotective function.

  18. Individual radiosensitivity does not correlate with radiation-induced apoptosis in lymphoblastoid cell lines or CD{sup 3+} lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Wistop, A.; Keller, U.; Grabenbauer, G.G.; Sauer, R.; Distel, L.V.R. [Dept. of Radiation Oncology, Friedrich Alexander Univ. Erlangen-Nuremberg, Erlangen (Germany); Sprung, C.N. [Div. of Research, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia)

    2005-05-01

    Background and purpose: spontaneous and radiation-induced apoptosis of lymphoblastoid cell lines (LCLs) derived from healthy donors, cancer patients and donors with radiosensitivity syndromes as well as CD{sup 3+} lymphocytes from patients with {>=} grade 3 late toxicity were investigated as a possible marker for the detection of individual radiosensitivity. These investigations are based on the hypothesis that hypersensitive patients have reduced levels of apoptosis after in vitro irradiation as a result of a defect in the signaling pathway. Material and methods: Epstein-Barr virus-(EBV-)transformed LCLs derived from five healthy donors, seven patients with heterozygous or homozygous genotype for ataxia-telangiectasia or Nijmegen breakage syndrome and five patients with {>=} grade 3 late toxicity (RTOG) were investigated. In addition, CD{sup 3+} lymphocytes from 21 healthy individuals and 18 cancer patients including five patients with a proven cellular hypersensitivity to radiation were analyzed. Cells were irradiated in vitro with a dose of 2 and 5 Gy and were incubated for 48 h. Apoptotic rates were measured by the TUNEL assay followed by customized image analysis. Results: four out of seven radiosensitivity syndrome patients were identified to have an increased cellular radiosensitivity as determined by reduced apoptotic rates after irradiation of their respective LCLs. Comparatively, only two of the five hypersensitive cancer patients were clearly identified by reduced apoptotic rates. Spontaneous apoptotic rates were very homogeneous among all 39 samples from controls and patients, while lymphocytes of all cancer patients showed significantly lower radiation-induced rates. Conclusion: only a subgroup of hypersensitive patients may be identified by reduction of radiation-induced apoptotic rate. It is concluded that the hypothesis according to which hypersensitive cells have reduced levels of apoptosis is only conditionally true. The authors suggest that this

  19. Newborn Screening for Primary T- and B-Cell Immune Deficiencies—A Prospective Study in Andalucía

    Directory of Open Access Journals (Sweden)

    Beatriz de Felipe

    2017-10-01

    Full Text Available Background: Quantification of T-cell-receptor-excision circles (TRECs and kappa-deleting-recombination-excision circles (KRECs from dried blood spots (DBS allows detection of neonates with severe T-cell and/or B-cell lymphopenia that are potentially affected by severe combined immunodeficiency (SCID, as well as X-linked agammaglobulinemia (XLA. Methods: Determination of TRECs and KRECs using a triplex RT-PCR (TRECS-KRECS-β-actin assay from prospectively collected DBS between February 2014 and December 2016 in three hospitals in Seville, Spain. Cut-off levels were TRECs < 6/punch, KRECs < 4/punch and b-actin > 700/punch. Internal (SCID, XLA, ataxia telangiectasia and external controls (CDC were included. Results: A total of 8943 DBS samples obtained from 8814 neonates were analysed. Re-punching was necessary in 124 samples (1.4% due to insufficient β-actin values (<700 copies/punch. Preterm neonates (GA < 37 weeks and neonates with a BW < 2500 g showed significantly lower TRECs and KRECs levels (p < 0.001. Due to repeated pathological results, ten neonates were re-sampled (0.11%, of which five neonates (0.055% confirmed the pathological results: one case was a fatal chromosomopathy (TRECs 1/KRECs 4; two were extreme premature newborns (TRECs 0/KRECs 0 and TRECs 1/KRECs 20 copies/punch; and 2 neonates were born to mothers receiving azathioprine during pregnancy (TRECs 92/KRECs 1 and TRECs 154/KRECs 3 copies/punch. All controls were correctly identified. Conclusions: Severe T- and B-cell lymphopenias were correctly identified by the TRECS-KRECS-β-actin assay. Prematurity and low BW are associated with lower TREC and KREC levels. Extreme prematurity and maternal immune suppressive therapy can cause false positive results of TRECs and KRECs values.

  20. N-Hydroxycinnamide derivatives of osthole presenting genotoxicity and cytotoxicity against human colon adenocarcinoma cells in vitro and in vivo.

    Science.gov (United States)

    Liu, Ling-Yu; Huang, Wei-Jan; Lin, Ren-Jye; Lin, Shyr-Yi; Liang, Yu-Chih

    2013-11-18

    Osthole is extracted from the Chinese herbs Cnidium monnieri and Angelica pubescens, and it was found to have antitumor activity in vitro and in vivo. A series of osthole derivatives have been synthesized, and the N-hydroxycinnamide derivatives of osthole, WJ1376-1 and WJ1398-1 were found to have the greatest potential against human colon adenocarcinoma cells. In contrast to the parental osthole, both WJ1376-1 and WJ1398-1 were found to induce multinucleation and polyploidy by microscopic observation and flow cytometry. WJ1376-1 and WJ1398-1 significantly activated ataxia telangiectasia and rad3 related (ATR) kinase, which triggered activation of the checkpoint kinase 2 (Chk2) signaling pathway and then down regulated Cdc25 phosphatase and Cdc2/cyclin B kinase activities. WJ1376-1 and WJ1398-1 also inhibited the phosphorylation of Aurora A kinase, which is associated with important processes during mitosis. The presence of a "comet" DNA fragment and phosphorylation of p53 at Ser 15 clearly indicated that DNA damage occurred with WJ1376-1 and WJ1398-1 treatment. WJ1376-1 and WJ1398-1 ultimately induced apoptosis as evidenced by the upregulation of Bad and activation of caspases-3, -7, and -9. Furthermore, WJ1376-1 and WJ1398-1 also showed a great effect in attenuating tumor growth without affecting the body weight of xenograft nude mice. Taken together, these results suggest that the toxic activities of WJ1376-1 and WJ1398-1 were dissimilar to that of the parental osthole, which can induce cell polyploidy and G2/M cell cycle arrest in colon adenocarcinoma cells and may provide a potential therapeutic target for colon cancer treatment in the future.

  1. Increased frequency of spontaneous neoplastic transformation in progeny of bystander cells from cultures exposed to densely ionizing radiation.

    Directory of Open Access Journals (Sweden)

    Manuela Buonanno

    Full Text Available An increased risk of carcinogenesis caused by exposure to space radiation during prolonged space travel is a limiting factor for human space exploration. Typically, astronauts are exposed to low fluences of ionizing particles that target only a few cells in a tissue at any one time. The propagation of stressful effects from irradiated to neighboring bystander cells and their transmission to progeny cells would be of importance in estimates of the health risks of exposure to space radiation. With relevance to the risk of carcinogenesis, we investigated, in model C3H 10T½ mouse embryo fibroblasts (MEFs, modulation of the spontaneous frequency of neoplastic transformation in the progeny of bystander MEFs that had been in co-culture 10 population doublings earlier with MEFs exposed to moderate doses of densely ionizing iron ions (1 GeV/nucleon or sparsely ionizing protons (1 GeV. An increase (P<0.05 in neoplastic transformation frequency, likely mediated by intercellular communication through gap junctions, was observed in the progeny of bystander cells that had been in co-culture with cells irradiated with iron ions, but not with protons.

  2. Argon protects hypoxia-, cisplatin- and gentamycin-exposed hair cells in the newborn rat's organ of Corti.

    Science.gov (United States)

    Yarin, Yury M; Amarjargal, Nyamaa; Fuchs, Julia; Haupt, Heidemarie; Mazurek, Birgit; Morozova, Svetlana V; Gross, Johann

    2005-03-01

    During the last few years, an important protective effect of the noble gas xenon against neuronal hypoxic damage was observed. However, argon (Ar), a gas from the same chemical group, but less expensive and without anesthetic effect at normobaric pressure, has not been studied in terms of possible biological effects on cell protection. Ar was tested for its ability to protect organotypic cultures of the organ of Corti from 3-5 day old rats against hypoxia, cisplatin, and gentamycin toxicity. Cultures were exposed to nitrogen hypoxia (5% CO2, 95% N2), Ar hypoxia (5% CO2, 95% Ar) or normoxia for 30 h. Ar protected the hair cells from hypoxia-induced damage by about 25%. Ar-oxygen (O2) mixtures (21% O2, 5% CO2, 74% Ar) had no effect on the hair cell survival. Cisplatin (7.5-25 microM) and gentamycin (5-40 microM) exposed in medium under air damaged the hair cells in a dose-dependent manner. The exposure of cisplatin- and gentamycin-treated cultures to the Ar-O2 atmosphere significantly reduced the hair cell damage by up to 25%. This protective effect of Ar might provide a new protective approach against ototoxic processes.

  3. Aryl Hydrocarbon Receptor Suppresses the Prostate Cancer LNCaP Cell Growth and Invasion by Promoting DNA Damage Response Under Oxidative Stress.

    Science.gov (United States)

    Yu, Jing-Song; Leng, Peng-Fei; Li, Yi-Fu; Wang, Yong-Quan; Wang, Yan; An, Rui-Hua; Qi, Ji-Ping

    2017-11-01

    Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that interacts with multiple signaling pathways during prostate development. In the present study, LNCaP cells were knocked down of AhR by siRNA, or treated with the AhR agonist 3-methylcholanthrene (3MC). The effects of AhR on LNCaP cells and the associated mechanisms were studied both under normal condition and under hydrogen peroxide (H 2 O 2 ) - induced oxidative stress. MTT, transwell chamber assays and flow cytometry were employed to investigate cell proliferation, invasion, and apoptosis, respectively, whereas the DNA damage response (DDR) signaling (phosphorylation of ataxia-telangiectasia mutated [ATM], check-point kinase 2 [Chk2], histone H2AX, p53, and cleaved poly-ADP-ribose polymerase [PARP]) was detected by western blotting. Exposure of LNCaP cells to H 2 O 2 inhibited their viability and migration, and induced apoptosis, at a greater extent compared with the culture under normal conditions. In addition, the oxidative stress increased p-ATM, p-Chk2, p-p53, and p-H2AX expression levels significantly. Knockdown of AhR attenuated the aforementioned effects caused by H 2 O 2 -induced oxidative stress. Activation of AhR by 3MC treatment, further aggravated these changes of LNCaP cells on oxidative stress. The findings indicated that AhR suppresses the viability and migration of LNCaP cells notably under oxidative stress, and this process is associated with positive regulation of the responses to oxidative DNA damage.

  4. Oxidative stress and apoptosis induction in human thyroid carcinoma cells exposed to the essential oil from Pistacia lentiscus aerial parts.

    Directory of Open Access Journals (Sweden)

    Simona Catalani

    Full Text Available Essential oils from the aerial parts (leaves, twigs and berries of Pistacia lentiscus (PLEO have been well characterized for their antibacterial and anti-inflammatory properties; however, poor information exists on their potential anticancer activity.Increasing concentrations of PLEO (0.01-0.1% v/v, 80-800 μg/ml were administered to a wide variety of cultured cancer cells from breast, cervix, colon, liver, lung, prostate, and thyroid carcinomas. Fibroblasts were also included as healthy control cells. Cell viability was monitored by WST-8 assay up to 72 hours after PLEO administration. The intracellular formation of reactive oxygen species (ROS, the induction of apoptosis, and the enhancement of chemotherapeutic drug cytotoxicity by PLEO were further investigated in the most responsive cancer cell line.A dose-dependent reduction of tumor cell viability was observed upon PLEO exposure; while no cytotoxic effect was revealed in healthy fibroblasts. FTC-133 thyroid cancer cells were found to be the most sensitive cells to PLEO treatment; accordingly, an intracellular accumulation of ROS and an activation of both the extrinsic and intrinsic apoptotic pathways were evidenced in FTC-133 cells after PLEO administration. Furthermore, the cytotoxic effect of the antineoplastic drugs cisplatin, 5-fluorouracil and etoposide was enhanced in PLEO-exposed FTC-133 cells.Taking into account its mode of action, PLEO might be considered as a promising source of natural antitumor agents which might have therapeutic potential in integrated oncology.

  5. Oxidative stress and apoptosis induction in human thyroid carcinoma cells exposed to the essential oil from Pistacia lentiscus aerial parts.

    Science.gov (United States)

    Catalani, Simona; Palma, Francesco; Battistelli, Serafina; Benedetti, Serena

    2017-01-01

    Essential oils from the aerial parts (leaves, twigs and berries) of Pistacia lentiscus (PLEO) have been well characterized for their antibacterial and anti-inflammatory properties; however, poor information exists on their potential anticancer activity. Increasing concentrations of PLEO (0.01-0.1% v/v, 80-800 μg/ml) were administered to a wide variety of cultured cancer cells from breast, cervix, colon, liver, lung, prostate, and thyroid carcinomas. Fibroblasts were also included as healthy control cells. Cell viability was monitored by WST-8 assay up to 72 hours after PLEO administration. The intracellular formation of reactive oxygen species (ROS), the induction of apoptosis, and the enhancement of chemotherapeutic drug cytotoxicity by PLEO were further investigated in the most responsive cancer cell line. A dose-dependent reduction of tumor cell viability was observed upon PLEO exposure; while no cytotoxic effect was revealed in healthy fibroblasts. FTC-133 thyroid cancer cells were found to be the most sensitive cells to PLEO treatment; accordingly, an intracellular accumulation of ROS and an activation of both the extrinsic and intrinsic apoptotic pathways were evidenced in FTC-133 cells after PLEO administration. Furthermore, the cytotoxic effect of the antineoplastic drugs cisplatin, 5-fluorouracil and etoposide was enhanced in PLEO-exposed FTC-133 cells. Taking into account its mode of action, PLEO might be considered as a promising source of natural antitumor agents which might have therapeutic potential in integrated oncology.

  6. CtIP-dependent DNA resection is required for DNA damage checkpoint maintenance but not initiation

    DEFF Research Database (Denmark)

    Kousholt, Arne Nedergaard; Fugger, Kasper; Hoffmann, Saskia

    2012-01-01

    To prevent accumulation of mutations, cells respond to DNA lesions by blocking cell cycle progression and initiating DNA repair. Homology-directed repair of DNA breaks requires CtIP-dependent resection of the DNA ends, which is thought to play a key role in activation of ATR (ataxia telangiectasia...

  7. Sirolimus Increases T-Cell Abundance in the Sun Exposed Skin of Kidney Transplant Recipients

    Directory of Open Access Journals (Sweden)

    Michael Thomas Burke, MBBS

    2017-07-01

    Conclusions. This study demonstrated that immunosuppressive drug class and sun exposure modify the abundance of multiple T-cell subsets in the skin of KTRs. Correlation analysis revealed that the prevalence of Treg cells in KTR blood does not accurately reflect the prevalence of Treg cells in KTR skin.

  8. Gli1 protein regulates the S-phase checkpoint in tumor cells via Bid protein, and its inhibition sensitizes to DNA topoisomerase 1 inhibitors.

    Science.gov (United States)

    Tripathi, Kaushlendra; Mani, Chinnadurai; Barnett, Reagan; Nalluri, Sriram; Bachaboina, Lavanya; Rocconi, Rodney P; Athar, Mohammed; Owen, Laurie B; Palle, Komaraiah

    2014-11-07

    Aberrant expression of hedgehog molecules, particularly Gli1, is common in cancers of many tissues and is responsible for their aggressive behavior and chemoresistance. Here we demonstrate a novel and tumor-specific role for aberrant Gli1 in the regulation of the S-phase checkpoint that suppresses replication stress and resistance to chemotherapy. Inhibition of Gli1 in tumor cells induced replication stress-mediated DNA damage response, attenuated their clonogenic potential, abrogated camptothecin (CPT)-induced Chk1 phosphorylation, and potentiated its cytotoxicity. However, in normal fibroblasts, Gli1 siRNAs showed no significant changes in CPT-induced Chk1 phosphorylation. Further analysis of ataxia telangiectasia and Rad3-related protein (ATR)/Chk1 signaling cascade genes in tumor cells revealed an unexpected mechanism whereby Gli1 regulates ATR-mediated Chk1 phosphorylation by transcriptional regulation of the BH3-only protein Bid. Consistent with its role in DNA damage response, Bid down-regulation in tumor cells abolished CPT-induced Chk1 phosphorylation and sensitized them to CPT. Correspondingly, Gli1 inhibition affected the expression of Bid and the association of replication protein A (RPA) with the ATR- interacting protein (ATRIP)-ATR complex, and this compromised the S-phase checkpoint. Conversely, complementation of Bid in Gli1-deficient cells restored CPT-induced Chk1 phosphorylation. An in silico analysis of the Bid promoter identified a putative Gli1 binding site, and further studies using luciferase reporter assays confirmed Gli1-dependent promoter activity. Collectively, our studies established a novel connection between aberrant Gli1 and Bid in the survival of tumor cells and their response to chemotherapy, at least in part, by regulating the S-phase checkpoint. Importantly, our data suggest a novel drug combination of Gli1 and Top1 inhibitors as an effective therapeutic strategy in treating tumors that expresses Gli1. © 2014 by The American

  9. Gli1 Protein Regulates the S-phase Checkpoint in Tumor Cells via Bid Protein, and Its Inhibition Sensitizes to DNA Topoisomerase 1 Inhibitors*

    Science.gov (United States)

    Tripathi, Kaushlendra; Mani, Chinnadurai; Barnett, Reagan; Nalluri, Sriram; Bachaboina, Lavanya; Rocconi, Rodney P.; Athar, Mohammed; Owen, Laurie B.; Palle, Komaraiah

    2014-01-01

    Aberrant expression of hedgehog molecules, particularly Gli1, is common in cancers of many tissues and is responsible for their aggressive behavior and chemoresistance. Here we demonstrate a novel and tumor-specific role for aberrant Gli1 in the regulation of the S-phase checkpoint that suppresses replication stress and resistance to chemotherapy. Inhibition of Gli1 in tumor cells induced replication stress-mediated DNA damage response, attenuated their clonogenic potential, abrogated camptothecin (CPT)-induced Chk1 phosphorylation, and potentiated its cytotoxicity. However, in normal fibroblasts, Gli1 siRNAs showed no significant changes in CPT-induced Chk1 phosphorylation. Further analysis of ataxia telangiectasia and Rad3-related protein (ATR)/Chk1 signaling cascade genes in tumor cells revealed an unexpected mechanism whereby Gli1 regulates ATR-mediated Chk1 phosphorylation by transcriptional regulation of the BH3-only protein Bid. Consistent with its role in DNA damage response, Bid down-regulation in tumor cells abolished CPT-induced Chk1 phosphorylation and sensitized them to CPT. Correspondingly, Gli1 inhibition affected the expression of Bid and the association of replication protein A (RPA) with the ATR- interacting protein (ATRIP)-ATR complex, and this compromised the S-phase checkpoint. Conversely, complementation of Bid in Gli1-deficient cells restored CPT-induced Chk1 phosphorylation. An in silico analysis of the Bid promoter identified a putative Gli1 binding site, and further studies using luciferase reporter assays confirmed Gli1-dependent promoter activity. Collectively, our studies established a novel connection between aberrant Gli1 and Bid in the survival of tumor cells and their response to chemotherapy, at least in part, by regulating the S-phase checkpoint. Importantly, our data suggest a novel drug combination of Gli1 and Top1 inhibitors as an effective therapeutic strategy in treating tumors that expresses Gli1. PMID:25253693

  10. Comparison of photovoltaic cell temperatures in modules operating with exposed and enclosed back surfaces

    Science.gov (United States)

    Namkoong, D.; Simon, F. F.

    1981-01-01

    Four different photovoltaic module designs were tested to determine the cell temperature of each design. The cell temperatures were compared to those obtained on identical design, using the same nominal operating cell temperature (NOCT) concept. The results showed that the NOCT procedure does not apply to the enclosed configurations due to continuous transient conditions. The enclosed modules had higher cell temperatures than the open modules, and insulated modules higher than the uninsulated. The severest performance loss - when translated from cell temperatures - 17.5 % for one enclosed, insulated module as a compared to that module mounted openly.

  11. Effect of amygdalin on the proliferation of hyperoxia-exposed type II alveolar epithelial cells isolated from premature rat.

    Science.gov (United States)

    Zhu, Huaping; Chang, Liwen; Li, Wenbin; Liu, Hanchu

    2004-01-01

    The pathogenesis of hyperoxia lung injury and the mechanism of amygdalin on type 2 alveolar epithelial cells (AEC2) isolated from premature rat lungs in vitro were investigated. AEC2 were obtained by primary culture from 20-days fetal rat lung and hyperoxia-exposed cell model was established. Cell proliferating viability was examined by MTT assay after treatment of amygdalin at various concentrations. DNA content and the proliferating cell nuclear antigen (PCNA) protein expression of AEC2 were measured by using flow cytometry and immunocytochemistry respectively after 24 h of hyperoxia exposure or amygdalin treatment. The results showed that hyperoxia inhibited the proliferation and decreased PCNA protein expression in A-EC2 of premature rat in vitro. Amygdalin at the concentration range of 50-200 micromol/L stimulated the proliferation of AEC2 in a dose-dependent manner, however, 400 micromol/L amygdalin inhibited the proliferation of AEC2. Amygdalin at the concentration of 200 micromol/L played its best role in facilitating proliferation of AEC2s in vitro and could partially ameliorated the changes of proliferation in hyperoxia exposed AEC2 of premature rat. It has been suggested that hyperoxia inhibited the proliferation of AEC2s of premature rat, which may contribute to hyperoxia lung injury. Amygdalin may play partial protective role in hyperoxia-induced lung injury.

  12. A DP based scheme for real-time reconfiguration of solar cell arrays exposed to dynamic changing inhomogeneous illuminations

    DEFF Research Database (Denmark)

    Shi, Liping; Brehm, Robert

    2016-01-01

    The overall energy conversion efficiency of solar cell arrays is highly effected by partial shading effects. Especially for solar panel arrays installed in environments which are exposed to inhomogeneous dynamic changing illuminations such as on roof tops of electrical vehicles the overall system...... efficiency is drastically reduced. Dynamic real-time reconfiguration of the solar panel array can reduce effects on the output efficiency due to partial shading. This results in a maximized power output of the panel array when exposed to dynamic changing illuminations. The optimal array configuration...... with respect to shading patterns can be stated as a combinatorial optimization problem and this paper proposes a dynamic programming (DP) based algorithm which finds the optimal feasible solution to reconfigure the solar panel array for maximum efficiency in real-time with linear time complexity. It is shown...

  13. A nitroimidazole derivative, PR-350, enhances the killing of pancreatic cancer cells exposed to high-dose irradiation under hypoxia

    Energy Technology Data Exchange (ETDEWEB)

    Mizumoto, Kazuhiro; Qian, Li-Wu; Zhang, Li.; Nagai, Eishi; Kura, Shinobu; Tanaka, Masao [Kyushu Univ., Fukuoka (Japan). Graduate School of Medical Sciences

    2002-03-01

    The radiosensitizing effects of PR-350, a nitroimidazole derivative, were examined concerning the cell killing of human pancreatic cancer cell lines exposed to high doses of gamma-ray irradiation in vitro. The percentages of dead cells were analyzed with a multiwell plate reader to measure the fluorescence intensity of propidium iodide before and after a digitonin treatment. The sensitizing effect of PR-350 on cell killing by high-dose irradiation was confirmed by time-course, dose-dependency, and microscopic observations. In five of seven pancreatic cancer cell lines in which the number of dead cells was determined 5 days after 30 Gy irradiation in the presence of PR-350, the number was significantly increased under hypoxic conditions, but not under aerobic conditions. The selective radiosensitive effect of PR-350 on hypoxic cells was also confirmed by flow cytometry. The results indicate that PR-350 can enhance the killing of pancreatic cancer cells by high-dose irradiation under hypoxia, which supports its clinical radiosensitizing effects when administered during intraoperative irradiation to pancreatic cancer. (author)

  14. Gene expression profiling and pathway analysis of human bronchial epithelial cells exposed to airborne particulate matter collected from Saudi Arabia

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Hong [Department of Environmental Medicine, NYU School of Medicine, Tuxedo, NY (United States); Shamy, Magdy [Department of Environmental Sciences, Faculty of Meteorology, Environment and Arid Land Agriculture, King Abdulaziz University, Jeddah (Saudi Arabia); Kluz, Thomas; Muñoz, Alexandra B.; Zhong, Mianhua; Laulicht, Freda [Department of Environmental Medicine, NYU School of Medicine, Tuxedo, NY (United States); Alghamdi, Mansour A.; Khoder, Mamdouh I. [Department of Environmental Sciences, Faculty of Meteorology, Environment and Arid Land Agriculture, King Abdulaziz University, Jeddah (Saudi Arabia); Chen, Lung-Chi [Department of Environmental Medicine, NYU School of Medicine, Tuxedo, NY (United States); Costa, Max, E-mail: Max.Costa@nyumc.org [Department of Environmental Medicine, NYU School of Medicine, Tuxedo, NY (United States)

    2012-12-01

    Epidemiological studies have established a positive correlation between human mortality and increased concentration of airborne particulate matters (PM). However, the mechanisms underlying PM related human diseases, as well as the molecules and pathways mediating the cellular response to PM, are not fully understood. This study aims to investigate the global gene expression changes in human cells exposed to PM{sub 10} and to identify genes and pathways that may contribute to PM related adverse health effects. Human bronchial epithelial cells were exposed to PM{sub 10} collected from Saudi Arabia for 1 or 4 days, and whole transcript expression was profiled using the GeneChip human gene 1.0 ST array. A total of 140 and 230 genes were identified that significantly changed more than 1.5 fold after PM{sub 10} exposure for 1 or 4 days, respectively. Ingenuity Pathway Analysis revealed that different exposure durations triggered distinct pathways. Genes involved in NRF2-mediated response to oxidative stress were up-regulated after 1 day exposure. In contrast, cells exposed for 4 days exhibited significant changes in genes related to cholesterol and lipid synthesis pathways. These observed changes in cellular oxidative stress and lipid synthesis might contribute to PM related respiratory and cardiovascular disease. -- Highlights: ► PM exposure modulated gene expression and associated pathways in BEAS-2B cells. ► One-day exposure to PM induced genes involved in responding to oxidative stress. ► 4-day exposure to PM changed genes associated to cholesterol and lipid synthesis.

  15. Inhibition of p21-activated kinase 4 expression suppresses the proliferation of Hep-2 laryngeal carcinoma cells via activation of the ATM/Chk1/2/p53 pathway.

    Science.gov (United States)

    Sun, Xin; Liu, Bin; Wang, Jin; Li, Jun; Ji, Wen-Yue

    2013-02-01

    In the present study, we investigated the effects of the p21-activated kinase 4 (Pak4) gene on Hep-2 laryngeal carcinoma cells in vivo and in vitro. The expression of Pak4 was downregulated using small interfering RNA (siRNA). The downregulation of Pak4 decreased the proliferation and increased apoptosis and S phase arrest in Hep-2 cells in vitro. In further experiments, we determined that the S/G(2) transition was obstructed by the downregulation of Pak4 using 5‑chloro-2'‑deoxyuridine (CldU) and 5‑iodo‑2'‑deoxyuridine (IdU) double staining. A xenografted Hep-2 tumor mouse model was created by inducing human tumors with a subcutaneous (s.c.) injection of 5x10(6) Hep-2 cells into the dorsal flank region of nu/nu mice. The downregulation of Pak4 in established xenografted tumors decreased tumor size and weight. The survival rate of the mice with tumors that did not express Pak4 was significantly higher compared to the mice with tumors expressing Pak4. These results confirm the role of Pak4 as an oncogene in laryngeal carcinoma cells. To identify the mechanism of the cell cycle arrest induced by Pak4, immunohistochemical staining was performed to detect changes in cell cycle‑related proteins. The results demonstrated that p53 was activated following the downregulation of Pak4. The levels of ataxia telangiectasia mutated (ATM), the upstream protein of checkpoint kinase (Chk)1 and Chk2, also increased. Therefore, we confirmed that the mechanisms of the Pak4-induced cell cycle arrest invovlve the activation of the ATM/Chk1/2/p53 pathway. These results may prove helpful for the development of novel therapies for the treatment of laryngeal carcinoma.

  16. Morphological alterations of Vero cell exposed to coplanar PCB 126 and noncoplanar PCB 153.

    Science.gov (United States)

    Shen, Kaili; Shen, Chaofeng; Chen, Lei; Chen, Xincai; Chen, Yingxu

    2012-01-01

    Polychlorinated biphenyls (PCBs) are widespread, persistent environmental contaminants that display a complex spectrum of toxicological properties. Exposure to PCBs has been associated with morphological anomalies in cell cultures. However, most mechanistic studies of PCBs' toxic activity have been focused on coplanar congeners. It is of importance to determine whether PCB treatment would influence cell configuration and whether these changes would depend on the structural characteristics of PCBs. In this study, we investigated cell morphological alteration in Vero cell cultures after exposure to coplanar PCB 126 and noncoplanar PCB 153. The survival of Vero cells was measured through the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test. Cytotoxicity results suggested that PCB congeners had a toxic, antiproliferative effect on Vero cells. Morphological studies described structural modifications and provided evidence that apoptosis might be the main cell death pathway in PCB 153-treated cells. The comparison between PCB 126 and PCB 153 indicated that the cell death mechanisms involved in coplanar or noncoplanar PCB congener exposure were different in Vero cells. Copyright © 2010 Wiley Periodicals, Inc.

  17. Glutaraldehyde fixation preserves the trend of elasticity alterations for endothelial cells exposed to TNF-α.

    Science.gov (United States)

    Targosz-Korecka, Marta; Brzezinka, Grzegorz Daniel; Danilkiewicz, Joanna; Rajfur, Zenon; Szymonski, Marek

    2015-03-01

    Among the users of atomic force microscopy based techniques, there is an ongoing discussion, whether cell elasticity measurements performed on fixed cells could be used for determination of the relative elasticity changes of the native (unfixed) cells subjected to physiologically active external agents. In this article, we present a case, for which the legitimacy of cell fixation for elasticity measurements is justified. We provide an evidence that the alterations of cell elasticity triggered by tumor necrosis factor alpha (TNF-α) in EA.hy926 endothelial cells are preserved after glutaraldehyde (GA) fixation. The value of post-fixation elasticity parameter is a product of the elasticity parameter obtained for living cells and a constant value, dependent on the GA concentration. The modification of the initial value of elasticity parameter caused by remodeling of the cortical actin cytoskeleton is reflected in the elasticity measurements performed on fixed cells. Thus, such fixation procedure may be particularly helpful for experiments, where the influence of an external agent on the cell cortex should be assessed and AFM measurements of living cells are problematic or better statistics is needed. © 2015 Wiley Periodicals, Inc.

  18. Area Expansivity Moduli of Regenerating Plant Protoplast Cell Walls Exposed to Shear Flows

    Science.gov (United States)

    Fujimura, Yuu; Iino, Masaaki; Watanabe, Ugai

    2005-05-01

    To control the elasticity of the plant cell wall, protoplasts isolated from cultured Catharanthus roseus cells were regenerated in shear flows of 115 s-1 (high shear) and 19.2 s-1 (low shear, as a control). The surface area expansivity modulus and the surface breaking strength of these regenerating protoplasts were measured by a micropipette aspiration technique. Cell wall synthesis was also measured using a cell wall-specific fluorescent dye. High shear exposure for 3 h doubled both the surface area modulus and breaking strength observed under low shear, significantly decreased cell wall synthesis, and roughly quadrupled the moduli of the cell wall. Based on the cell wall synthesis data, we estimated the three-dimensional modulus of the cell wall to be 4.1± 1.2 GPa for the high shear, and 0.35± 0.2 GPa for the low shear condition, using the surface area expansivity modulus divided by the cell wall thickness, which is identical with the Young’s modulus divided by 2(1-σ), where σ is Poisson's ratio. We concluded that high shear exposure considerably strengthens the newly synthesized cell wall.

  19. Structural damage of chicken red blood cells exposed to platinum nanoparticles and cisplatin

    DEFF Research Database (Denmark)

    Kutwin, Marta; Sawosz, Ewa; Jaworski, Sławomir

    2014-01-01

    of platinum nanoparticles (NP-Pt) and cisplatin with blood compartments are important for future applications. This study investigated structural damage, cell membrane deformation and haemolysis of chicken embryo red blood cells (RBC) after treatment with cisplatin and NP-Pt. Cisplatin (4 μg/ml) and NP-Pt (2......,6 μg/ml), when incubated with chicken embryo RBC, were detrimental to cell structure and induced haemolysis. The level of haemolytic injury was increased after cisplatin and NP-Pt treatments compared to the control group. Treatment with cisplatin caused structural damage to cell membranes...... and the appearance of keratocytes, while NP-Pt caused cell membrane deformations (discoid shape of cells was lost) and the formation of knizocytes and echinocytes. This work demonstrated that NP-Pt have potential applications in anticancer therapy, but potential toxic side effects must be explored in future...

  20. Genotoxicity assessment data for exfoliated buccal cells exposed to mobile phone radiation

    Directory of Open Access Journals (Sweden)

    F.M. de Oliveira

    2017-12-01

    Full Text Available Healthy mobile phone users aged 18–30 y.o. provided exfoliated buccal cells samples from the right and left inner cheeks. A total of 2000 cells per subject were screened for the presence of micronuclei as a sign of genotoxic damage, according to the mobile phone use profile of each user. Keywords: Electromagnetic fields, Mobile phones, Genotoxicity, Micronuclei, Exfoliated buccal cells, Feulgen stain

  1. ATP Release from Human Airway Epithelial Cells Exposed to Staphylococcus aureus Alpha-Toxin

    Directory of Open Access Journals (Sweden)

    Romina Baaske

    2016-12-01

    Full Text Available Airway epithelial cells reduce cytosolic ATP content in response to treatment with S. aureus alpha-toxin (hemolysin A, Hla. This study was undertaken to investigate whether this is due to attenuated ATP generation or to release of ATP from the cytosol and extracellular ATP degradation by ecto-enzymes. Exposure of cells to rHla did result in mitochondrial calcium uptake and a moderate decline in mitochondrial membrane potential, indicating that ATP regeneration may have been attenuated. In addition, ATP may have left the cells through transmembrane pores formed by the toxin or through endogenous release channels (e.g., pannexins activated by cellular stress imposed on the cells by toxin exposure. Exposure of cells to an alpha-toxin mutant (H35L, which attaches to the host cell membrane but does not form transmembrane pores, did not induce ATP release from the cells. The Hla-mediated ATP-release was completely blocked by IB201, a cyclodextrin-inhibitor of the alpha-toxin pore, but was not at all affected by inhibitors of pannexin channels. These results indicate that, while exposure of cells to rHla may somewhat reduce ATP production and cellular ATP content, a portion of the remaining ATP is released to the extracellular space and degraded by ecto-enzymes. The release of ATP from the cells may occur directly through the transmembrane pores formed by alpha-toxin.

  2. Associations of ATM Polymorphisms With Survival in Advanced Esophageal Squamous Cell Carcinoma Patients Receiving Radiation Therapy

    Energy Technology Data Exchange (ETDEWEB)

    Du, Zhongli [State Key Laboratory of Molecular Oncology, Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Department of Etiology and Carcinogenesis (Beijing Key Laboratory for Carcinogenesis and Cancer Prevention), Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Zhang, Wencheng [Department of Radiation Oncology, Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Zhou, Yuling; Yu, Dianke; Chen, Xiabin; Chang, Jiang; Qiao, Yan; Zhang, Meng; Huang, Ying; Wu, Chen [State Key Laboratory of Molecular Oncology, Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Department of Etiology and Carcinogenesis (Beijing Key Laboratory for Carcinogenesis and Cancer Prevention), Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Xiao, Zefen, E-mail: xiaozefen@sina.com [Department of Radiation Oncology, Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Tan, Wen, E-mail: tanwen@cicams.ac.cn [State Key Laboratory of Molecular Oncology, Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); Department of Etiology and Carcinogenesis (Beijing Key Laboratory for Carcinogenesis and Cancer Prevention), Cancer Institute and Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing (China); and others

    2015-09-01

    Purpose: To investigate whether single nucleotide polymorphisms (SNPs) in the ataxia telangiectasia mutated (ATM) gene are associated with survival in patients with esophageal squamous cell carcinoma (ESCC) receiving radiation therapy or chemoradiation therapy or surgery only. Methods and Materials: Four tagSNPs of ATM were genotyped in 412 individuals with clinical stage III or IV ESCC receiving radiation therapy or chemoradiation therapy, and in 388 individuals with stage I, II, or III ESCC treated with surgery only. Overall survival time of ESCC among different genotypes was estimated by Kaplan-Meier plot, and the significance was examined by log-rank test. The hazard ratios (HRs) and 95% confidence intervals (CIs) for death from ESCC among different genotypes were computed by a Cox proportional regression model. Results: We found 2 SNPs, rs664143 and rs664677, associated with survival time of ESCC patients receiving radiation therapy. Individuals with the rs664143A allele had poorer median survival time compared with the rs664143G allele (14.0 vs 20.0 months), with the HR for death being 1.45 (95% CI 1.12-1.89). Individuals with the rs664677C allele also had worse median survival time than those with the rs664677T allele (14.0 vs 23.5 months), with the HR of 1.57 (95% CI 1.18-2.08). Stratified analysis showed that these associations were present in both stage III and IV cancer and different radiation therapy techniques. Significant associations were also found between the SNPs and locosregional progression or progression-free survival. No association between these SNPs and survival time was detected in ESCC patients treated with surgery only. Conclusion: These results suggest that the ATM polymorphisms might serve as independent biomarkers for predicting prognosis in ESCC patients receiving radiation therapy.

  3. Analysis of Pseudomonas aeruginosa cell envelope proteome by capture of surface-exposed proteins on activated magnetic nanoparticles.

    Directory of Open Access Journals (Sweden)

    Davide Vecchietti

    Full Text Available We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedded in the cell envelope fragments. For a high number of proteins, our analysis strongly indicates either surface exposure or localization in an envelope district. The localization of most identified proteins was only predicted or totally unknown. This novel approach greatly improves the sensitivity and specificity of the previous methods, such as surface shaving with proteases that was also tested on P. aeruginosa. The magneto-capture procedure is simple, safe, and rapid, and appears to be well-suited for envelope studies in highly pathogenic bacteria.

  4. Anti-Inflammatory Effects of Pomegranate Peel Extract in THP-1 Cells Exposed to Particulate Matter PM10

    Directory of Open Access Journals (Sweden)

    Soojin Park

    2016-01-01

    Full Text Available Epidemiological and experimental evidence support health risks associated with the exposure to airborne particulate matter with a diameter of <10 μM (PM10. PM10 stimulates the production of reactive oxygen species (ROS and inflammatory mediators. Thus, we assumed that natural antioxidants might provide health benefits attenuating hazardous effects of PM10. In the present study, we examined the effects of pomegranate peel extract (PPE on THP-1 monocytic cells exposed to PM10. PM10 induced cytotoxicity and the production of ROS. It also increased the expression and secretion of inflammatory cytokines, such as tumor necrosis factor-α (TNF-α, interleukin-1β (IL-1β, and monocyte chemoattractant protein-1 (MCP-1, and cell adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1. PPE at 10–100 μg mL−1 attenuated the production of ROS and the expression of TNF-α, IL-1β, MCP-1, and ICAM-1, but not VCAM-1, in THP-1 cells stimulated by PM10 (100 μg mL−1. PPE also attenuated the adhesion of PM10-stimulated THP-1 cells to EA.hy926 endothelial cells. PPE constituents, punicalagin and ellagic acid, attenuated PM10-induced monocyte adhesion to endothelial cells, and punicalagin was less cytotoxic compared to ellagic acid. The present study suggests that PPE and punicalagin may be useful in alleviating inflammatory reactions due to particulate matter.

  5. Protein thiol oxidation and formation of S-glutathionylated cyclophilin A in cells exposed to chloramines and hypochlorous acid.

    Science.gov (United States)

    Stacey, Melissa M; Cuddihy, Sarah L; Hampton, Mark B; Winterbourn, Christine C

    2012-11-01

    Neutrophil oxidants, including the myeloperoxidase products, HOCl and chloramines, have been linked to endothelial dysfunction in inflammatory diseases such as atherosclerosis. As they react preferentially with sulfur centers, thiol proteins are likely to be cellular targets. Our objectives were to establish whether there is selective protein oxidation in vascular endothelial cells treated with HOCl or chloramines, and to identify sensitive proteins. Cells were treated with HOCl, glycine chloramine and monochloramine, reversibly oxidized cysteines were labeled and separated by 1D or 2D SDS-PAGE, and proteins were characterized by mass spectrometry. Selective protein oxidation was observed, with chloramines and HOCl causing more changes than H(2)O(2). Cyclophilin A was one of the most sensitive targets, particularly with glycine chloramine. Cyclophilin A was also oxidized in Jurkat T cells where its identity was confirmed using a knockout cell line. The product was a mixed disulfide with glutathione, with glutathionylation at Cys-161. Glyceraldehyde-3-phosphate dehydrogenase, peroxiredoxins and cofilin were also highly sensitive to HOCl/chloramines. Cyclophilins are becoming recognized as redox regulatory proteins, and glutathionylation is an important mechanism for redox regulation. Cells lacking Cyclophilin A showed more glutathionylation of other proteins than wild-type cells, suggesting that cyclophilin-regulated deglutathionylation could contribute to redox changes in HOCl/chloramine-exposed cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Light-induced transpiration alters cell water relations in figleaf gourd (Cucurbita ficifolia) seedlings exposed to low root temperatures.

    Science.gov (United States)

    Lee, Seong Hee; Zwiazek, Janusz J; Chung, Gap Chae

    2008-06-01

    Water relation parameters including elastic modulus (epsilon), half-times of water exchange (T(w)(1/2)), hydraulic conductivity and turgor pressure (P) were measured in individual root cortical and cotyledon midrib cells in intact figleaf gourd (Cucurbita ficifolia) seedlings, using a cell pressure probe. Transpiration rates (E) of cotyledons were also measured using a steady-state porometer. The seedlings were exposed to low ambient (approximately 10 micromol m(-2) s(-1)) or high supplemental irradiance (approximately 300 micromol m(-2) s(-1) PPF density) at low (8 degrees C) or warm (22 degrees C) root temperatures. When exposed to low irradiance, all the water relation parameters of cortical cells remained similar at both root temperatures. The exposure of cotyledons to supplemental light at warm root temperatures, however, resulted in a two- to three-fold increase in T(w)(1/2) values accompanied with the reduced hydraulic conductivity in both root cortical (Lp) and cotyledon midrib cells (Lp(c)). Low root temperature (LRT) further reduced Lp(c) and E, whether it was measured under low or high irradiance levels. The reductions of Lp as the result of respective light and LRT treatments were prevented by the application of 1 microM ABA. Midrib cells required higher concentrations of ABA (2 microM) in order to prevent the reduction in Lp(c). When the exposure of cotyledons to light was accompanied by LRT, however, ABA proved ineffective in reversing the inhibition of Lp. LRT combined with high irradiance triggered a drastic 10-fold reduction in water permeability of cortical and midrib cells and increased epsilon and T(w)(1/2) values. Measurement of E indicated that the increased water demand by the transpiring plants was fulfilled by an increase in the apoplastic pathway as principal water flow route. The importance of water transport regulation by transpiration affecting the hydraulic conductivity of the roots is discussed.

  7. Naphthalimides Induce G2 Arrest Through the ATM-Activated Chk2-Executed Pathway in HCT116 Cells

    Directory of Open Access Journals (Sweden)

    Hong Zhu

    2009-11-01

    Full Text Available Naphthalimides, particularly amonafide and 2-(2-dimethylamino-6-thia-2-aza-benzo[def]chrysene-1,3-diones (R16, have been identified to possess anticancer activities and to induce G2-M arrest through inhibiting topoisomerase II accompanied by Chk1 degradation. The current study was designed to precisely dissect the signaling pathway(s responsible for the naphthalimide-induced cell cycle arrest in human colon carcinoma HCT116 cells. Using phosphorylated histone H3 and mitotic protein monoclonal 2 as mitosis markers, we first specified the G2 arrest elicited by the R16 and amonafide. Then, R16 and amonafide were revealed to induce phosphorylation of the DNA damage sensor ataxia telangiectasia-mutated (ATM responding to DNA double-strand breaks (DSBs. Inhibition of ATM by both the pharmacological inhibitor caffeine and the specific small interference RNA (siRNA rescued the G2 arrest elicited by R16, indicating its ATM-dependent characteristic. Furthermore, depletion of Chk2, but not Chk1 with their corresponding siRNA, statistically significantly reversed the R16- and amonafide-triggered G2 arrest. Moreover, the naphthalimides phosphorylated Chk2 in an ATM-dependent manner but induced Chk1 degradation. These data indicate that R16 and amonafide preferentially used Chk2 as evidenced by the differential ATM-executed phosphorylation of Chk1 and Chk2. Thus, a clear signaling pathway can be established, in which ATM relays the DNA DSBs signaling triggered by the naphthalimides to the checkpoint kinases, predominantly to Chk2,which finally elicits G2 arrest. The mechanistic elucidation not only favors the development of the naphthalimides as anticancer agents but also provides an alternative strategy of Chk2 inhibition to potentiate the anticancer activities of these agents.

  8. Dendritic cells inversely regulate airway inflammation in cigarette smoke-exposed mice

    NARCIS (Netherlands)

    Ezzati Givi, Masoumeh; Akbari, Peyman; Boon, Louis; Puzovic, Vladimir S; Bezemer, Gillina F G; Ricciardolo, Fabio L M; Folkerts, Gert; Redegeld, Frank A; Mortaz, Esmaeil

    The recruitment and activation of inflammatory cells into the respiratory system is considered a crucial feature in the pathophysiology of chronic obstructive pulmonary disease (COPD). Since dendritic cells (DCs) have a pivotal role in the onset and regulation of immune responses, we investigated

  9. Microcephalin and pericentrin regulate mitotic entry via centrosome-associated Chk1

    DEFF Research Database (Denmark)

    Tibelius, Alexandra; Marhold, Joachim; Zentgraf, Hanswalter

    2009-01-01

    in microcephalin (MCPH1), cells from patients with Seckel syndrome and MOPD II harbor mutations in ataxia telangiectasia and Rad3 related (ATR) or pericentrin (PCNT), leading to disturbed ATR signaling. In this study, we show that a lack of MCPH1 or PCNT results in a loss of Chk1 from centrosomes with subsequently...

  10. Loss of ATM kinase activity leads to embryonic lethality in mice

    DEFF Research Database (Denmark)

    Daniel, J.A.; Pellegrini, M.; Filsuf, D.

    2012-01-01

    Ataxia telangiectasia (A-T) mutated (ATM) is a key deoxyribonucleic acid (DNA) damage signaling kinase that regulates DNA repair, cell cycle checkpoints, and apoptosis. The majority of patients with A-T, a cancer-prone neurodegenerative disease, present with null mutations in Atm. To determine wh...

  11. Transcriptome analysis of the human T lymphocyte cell line Jurkat and human peripheral blood mononuclear cells exposed to deoxynivalenol (DON): New mechanistic insights

    Energy Technology Data Exchange (ETDEWEB)

    Katika, Madhumohan R. [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Hendriksen, Peter J.M. [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Shao, Jia [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Loveren, Henk van [Department of Health Risk Analysis and Toxicology, Maastricht University (Netherlands); National Institute for Public Health and the Environment (RIVM), Bilthoven (Netherlands); Netherlands Toxicogenomics Centre (Netherlands); Peijnenburg, Ad, E-mail: ad.peijnenburg@wur.nl [RIKILT-Institute of Food Safety, Wageningen University and Research Centre, Wageningen (Netherlands); Netherlands Toxicogenomics Centre (Netherlands)

    2012-10-01

    Deoxynivalenol (DON) or vomitoxin is a commonly encountered type-B trichothecene mycotoxin, produced by Fusarium species predominantly found in cereals and grains. DON is known to exert toxic effects on the gastrointestinal, reproductive and neuroendocrine systems, and particularly on the immune system. Depending on dose and exposure time, it can either stimulate or suppress immune function. The main objective of this study was to obtain a deeper insight into DON-induced effects on lymphoid cells. For this, we exposed the human T-lymphocyte cell line Jurkat and human peripheral blood mononuclear cells (PBMCs) to various concentrations of DON for various times and examined gene expression changes by DNA microarray analysis. Jurkat cells were exposed to 0.25 and 0.5 μM DON for 3, 6 and 24 h. Biological interpretation of the microarray data indicated that DON affects various processes in these cells: It upregulates genes involved in ribosome structure and function, RNA/protein synthesis and processing, endoplasmic reticulum (ER) stress, calcium-mediated signaling, mitochondrial function, oxidative stress, the NFAT and NF-κB/TNF-α pathways, T cell activation and apoptosis. The effects of DON on the expression of genes involved in ER stress, NFAT activation and apoptosis were confirmed by qRT-PCR. Other biochemical experiments confirmed that DON activates calcium-dependent proteins such as calcineurin and M-calpain that are known to be involved in T cell activation and apoptosis. Induction of T cell activation was also confirmed by demonstrating that DON activates NFATC1 and induces its translocation from the cytoplasm to the nucleus. For the gene expression profiling of PBMCs, cells were exposed to 2 and 4 μM DON for 6 and 24 h. Comparison of the Jurkat microarray data with those obtained with PBMCs showed that most of the processes affected by DON in the Jurkat cell line were also affected in the PBMCs. -- Highlights: ► The human T cell line Jurkat and human

  12. Whole-Genome Expression Analysis of Human Mesenchymal Stromal Cells Exposed to Ultrasmooth Tantalum vs. Titanium Oxide Surfaces

    DEFF Research Database (Denmark)

    Stiehler, C.; Bunger, C.; Overall, R. W.

    2013-01-01

    to Ti surface. Key genes related to osteogenesis and cell adhesion were upregulated by MSCs exposed to Ta. We further identified differentially regulated candidate transcription factors, e.g., NRF2, EGR1, IRF-1, IRF-8, NF-Y, and p53 as well as relevant signaling pathways, e.g., p53 and mTOR, indicating...... to titanium (Ti) surface. The aim of this study was to extend the previous investigation of biocompatibility by monitoring temporal gene expression of MSCs on topographically comparable smooth Ta and Ti surfaces using whole-genome gene expression analysis. Total RNA samples from telomerase-immortalized human...

  13. Low-level laser therapy: Effects on human face aged skin and cell viability of HeLa cells exposed to UV radiation

    Directory of Open Access Journals (Sweden)

    Mezghani Sana

    2015-01-01

    Full Text Available Chronic and excessive exposure to UV radiation leads to photoaging and photocarcinogenesis. Adequate protection of the skin against the deleterious effects of UV irradiation is essential. Low-level laser therapy (LLLT is a light source in the red to near-infrared range that has been accepted in a variety of medical applications. In this study, we explored the effect of LLLT in human face aged skin and the cell viability of HeLa cells exposed to UV radiation. We found that LLLT significantly reduced visible wrinkles and the loss of firmness of facial skin in aging subjects. Additionally, treatment of cultured HeLa cells with LLLT prior to or post UVA or UVB exposure significantly protected cells from UV-mediated cell death. All results showed the beneficial effects of LLLT on relieving signs of skin aging and its prevention and protection of the cell viability against UV-induced damage.

  14. Purinergic signaling mediates oxidative stress in UVA-exposed THP-1 cells

    Directory of Open Access Journals (Sweden)

    Ayumi Kawano

    2015-01-01

    Full Text Available Ultraviolet A (UVA radiation, the major UV component of solar radiation, can penetrate easily to the dermis, where it causes significant damage to cellular components by inducing formation of reactive oxygen species (ROS. On the other hand, extracellular ATP is released in response to various stimuli, and activates purinergic P2X7 receptor, triggering ROS production and cell death. Here, we examined the hypothesis that ATP release followed by activation of P2X7 receptor plays a role in UVA-induced oxidative cell damage, using human acute monocytic leukemia cell line THP-1. Indeed, UVA irradiation of THP-1 cells induced ATP release and activation of P2X7 receptor. Irradiated cells showed a rapid increase of both p67phox in membrane fraction and intracellular ROS. Pretreatment with ecto-nucleotidase or P2X7 receptor antagonist blocked the UVA-initiated membrane translocation of p67phox and ROS production. Furthermore, pretreatment with antioxidant or P2X7 receptor antagonist efficiently protected UVA-irradiated cells from caspase-dependent cell death. These findings show that autocrine signaling through release of ATP and activation of P2X7 receptor is required for UVA-induced stimulation of oxidative stress in monocytes.

  15. Structural damage of chicken red blood cells exposed to platinum nanoparticles and cisplatin

    Science.gov (United States)

    Kutwin, Marta; Sawosz, Ewa; Jaworski, Sławomir; Kurantowicz, Natalia; Strojny, Barbara; Chwalibog, André

    2014-05-01

    Side effects and resistance of cancer cells to cisplatin are major drawbacks to its application, and recently, the possibility of replacing cisplatin with nanocompounds has been considered. Most chemotherapeutic agents are administered intravenously, and comparisons between the interactions of platinum nanoparticles (NP-Pt) and cisplatin with blood compartments are important for future applications. This study investigated structural damage, cell membrane deformation and haemolysis of chicken embryo red blood cells (RBC) after treatment with cisplatin and NP-Pt. Cisplatin (4 μg/ml) and NP-Pt (2,6 μg/ml), when incubated with chicken embryo RBC, were detrimental to cell structure and induced haemolysis. The level of haemolytic injury was increased after cisplatin and NP-Pt treatments compared to the control group. Treatment with cisplatin caused structural damage to cell membranes and the appearance of keratocytes, while NP-Pt caused cell membrane deformations (discoid shape of cells was lost) and the formation of knizocytes and echinocytes. This work demonstrated that NP-Pt have potential applications in anticancer therapy, but potential toxic side effects must be explored in future preclinical research.

  16. Anatase TiO2 Nanoparticles with Exposed {001} Facets for Efficient Dye-Sensitized Solar Cells

    Science.gov (United States)

    Chu, Liang; Qin, Zhengfei; Yang, Jianping; Li, Xing’ao

    2015-01-01

    Anatase TiO2 nanoparticles with exposed {001} facets were synthesized from Ti powder via a sequential hydrothermal reaction process. At the first-step hydrothermal reaction, H-titanate nanowires were obtained in NaOH solution with Ti powder, and at second-step hydrothermal reaction, anatase TiO2 nanoparticles with exposed {001} facets were formed in NH4F solution. If the second-step hydrothermal reaction was carried out in pure water, the H-titanate nanowires were decomposed into random shape anatase-TiO2 nanostructures, as well as few impurity of H2Ti8O17 phase and rutile TiO2 phase. Then, the as-prepared TiO2 nanostructures synthesized in NH4F solution and pure water were applied to the photoanodes of dye-sensitized solar cells (DSSCs), which exhibited power conversion efficiency (PCE) of 7.06% (VOC of 0.756 V, JSC of 14.80 mA/cm2, FF of 0.631) and 3.47% (VOC of 0.764 V, JSC of 6.86 mA/cm2, FF of 0.662), respectively. The outstanding performance of DSSCs based on anatase TiO2 nanoparticles with exposed {001} facets was attributed to the high activity and large special surface area for excellent capacity of dye adsorption. PMID:26190140

  17. Regulation of SUMO2 Target Proteins by the Proteasome in Human Cells Exposed to Replication Stress

    DEFF Research Database (Denmark)

    Bursomanno, Sara; McGouran, Joanna F; Kessler, Benedikt M

    2015-01-01

    In human cells, SUMO2 is predominantly conjugated to target proteins in response to cellular stress. Previous studies suggested that proteins conjugated to SUMO2, but not to SUMO1, could be regulated by the ubiquitin-mediated proteasome system. Hence, we set out to understand the role...... of the proteasome in determining the fate of proteins conjugated to SUMO2 when cells are treated with DNA replication stress conditions. We conducted a quantitative proteomic analysis in a U2OS cell line stably expressing SUMO2(Q87R) tagged with StrepHA in the presence or absence of epoxomicin (EPOX), a proteasome...

  18. Effect of roflumilast on inflammatory cells in the lungs of cigarette smoke-exposed mice

    Directory of Open Access Journals (Sweden)

    De Cunto Giovanna

    2008-08-01

    Full Text Available Abstract Background We reported that roflumilast, a phosphodiesterase 4 inhibitor, given orally at 5 mg/kg to mice prevented the development of emphysema in a chronic model of cigarette smoke exposure, while at 1 mg/kg was ineffective. Here we investigated the effects of roflumilast on the volume density (VV of the inflammatory cells present in the lungs after chronic cigarette smoke exposure. Methods Slides were obtained from blocks of the previous study and VV was assessed immunohistochemically and by point counting using a grid with 48 points, a 20× objective and a computer screen for a final magnification of 580×. Neutrophils were marked with myeloperoxidase antibody, macrophages with Mac-3, dendritic cells with fascin, B-lymphocytes with B220, CD4+ T-cells with CD4+ antibody, and CD8+T-cells with CD8-α. The significance of the differences was calculated using one-way analysis of variance. Results Chronic smoke exposure increased neutrophil VV by 97%, macrophage by 107%, dendritic cell by 217%, B-lymphocyte by 436%, CD4+ by 524%, and CD8+ by 417%. The higher dose of roflumilast prevented the increase in neutrophil VV by 78%, macrophage by 82%, dendritic cell by 48%, B-lymphocyte by 100%, CD4+ by 98% and CD8+ VV by 88%. The lower dose of roflumilast did not prevent the increase in neutrophil, macrophage and B-cell VV but prevented dendritic cells by 42%, CD4+ by 55%, and CD8+ by 91%. Conclusion These results indicate (i chronic exposure to cigarette smoke in mice results in a significant recruitment into the lung of inflammatory cells of both the innate and adaptive immune system; (ii roflumilast at the higher dose exerts a protective effect against the recruitment of all these cells and at the lower dose against the recruitment of dendritic cells and T-lymphocytes; (iii these findings underline the role of innate immunity in the development of pulmonary emphysema and (iiii support previous results indicating that the inflammatory cells of

  19. Radioprotection by DMSO in nitrogen-saturated mammalian cells exposed to helium ion beams

    Science.gov (United States)

    Hirayama, Ryoichi; Matsumoto, Yoshitaka; Kase, Yuki; Noguchi, Miho; Ando, Koichi; Ito, Atsushi; Okayasu, Ryuichi; Furusawa, Yoshiya

    2009-12-01

    The contribution of OH radical-mediated indirect action by particle beams under hypoxic irradiation condition was investigated by using a radical scavenger. V79 cells were irradiated with 150 MeV/nucleon helium ions at an LET of 2.2 keV/μm in the presence or absence of DMSO, and their colony survivals were determined. The contribution of indirect action to cell killing under hypoxic condition was estimated to be 52±9%. We conclude that OH radical-mediated indirect action still has a half in total contribution on cell killing under hypoxic condition.

  20. Amperometric assessment of functional changes in nanoparticle-exposed immune cells: varying Au nanoparticle exposure time and concentration.

    Science.gov (United States)

    Marquis, Bryce J; Maurer-Jones, Melissa A; Braun, Katherine L; Haynes, Christy L

    2009-11-01

    A mast cell/fibroblast co-culture system is used as a model to assess the toxicity of Au nanoparticles over the course of 72 hours of exposure. Cellular uptake of nanoparticles was found to increase over the 72 hr exposure period and the nanoparticles localized within granular bodies of the primary culture mast cells. These granules were found to increase in volume with the addition of nanoparticles. There was no decrease in viability for 24 hr exposed cells but a slight viability decrease was found after 48 and 72 hr exposure. Carbon-fiber amperometry analysis of exocytosis of serotonin from mast cells revealed changing release profiles over the time course of exposure. In early exposure times, granular secretion of serotonin increased with exposure to Au nanoparticles whereas 72 hr exposure showed decreased secretion of serotonin with nanoparticle exposure. The kinetics of this release was also found to be affected by Au colloid exposure where the rate of serotonin expulsion decreased with increasing nanoparticle exposure. These results illustrate the dynamic nature of nanoparticle-cell interactions and the critical changes in cell behavior even when viability is unaffected.

  1. Nitric oxide measurements in hTERT-RPE cells and subcellular fractions exposed to low levels of red light

    Science.gov (United States)

    Wigle, Jeffrey C.; Castellanos, Cherry C.; Denton, Michael L.; Holwitt, Eric A.

    2014-02-01

    Cells in a tissue culture model for laser eye injury exhibit increased resistance to a lethal pulse of 2.0-μm laser radiation if the cells are first exposed to 2.88 J/cm2 of red light 24 hr prior to the lethal laser exposure. Changes in expression of various genes associated with apoptosis have been observed, but the biochemical link between light absorption and gene expression remains unknown. Cytochome c oxidase (CCOX), in the electron transport chain, is the currentlyhypothesized absorber. Absorption of the red light by CCOX is thought to facilitate displacement of nitric oxide (NO) by O2 in the active site, increasing cellular respiration and intracellular ATP. However, NO is also an important regulator and mediator of numerous physiological processes in a variety of cell and tissue types that is synthesized from l-arginine by NO synthases. In an effort to determine the relative NO contributions from these competing pathways, we measured NO levels in whole cells and subcellular fractions, with and without exposure to red light, using DAF-FM, a fluorescent dye that stoichiometrically reacts with NO. Red light induced a small, but consistently reproducible, increase in fluorescence intensity in whole cells and some subcellular fractions. Whole cells exhibited the highest overall fluorescence intensity followed by (in order) cytosolic proteins, microsomes, then nuclei and mitochondria.

  2. DNA damage and apoptosis of endometrial cells cause loss of the early embryo in mice exposed to carbon disulfide

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bingzhen [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China); Shen, Chunzi [Centers for Disease Control and Prevention, Zibo (China); Yang, Liu; Li, Chunhui; Yi, Anji [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China); Wang, Zhiping, E-mail: zhipingw@sdu.edu.cn [Department of Epidemiology and Health Statistics, School of Public Health, Shandong University, Jinan (China)

    2013-12-01

    Carbon disulfide (CS{sub 2}) may lead to spontaneous abortion and very early pregnancy loss in women exposed in the workplace, but the mechanism remains unclear. We designed an animal model in which gestating Kunming strain mice were exposed to CS{sub 2} via i.p. on gestational day 4 (GD4). We found that the number of implanted blastocysts on GD8 was significantly reduced by each dose of 0.1 LD{sub 50} (157.85 mg/kg), 0.2 LD{sub 50} (315.7 mg/kg) and 0.4 LD{sub 50} (631.4 mg/kg). In addition, both the level of DNA damage and apoptosis rates of endometrial cells on GD4.5 were increased, showed definite dose–response relationships, and inversely related to the number of implanted blastocysts. The expressions of mRNA and protein for the Bax and caspase-3 genes in the uterine tissues on GD4.5 were up-regulated, while the expressions of mRNA and protein for the Bcl-2 gene were dose-dependently down-regulated. Our results indicated that DNA damage and apoptosis of endometrial cells were important reasons for the loss of implanted blastocysts induced by CS{sub 2}. - Highlights: • We built an animal model of CS2 exposure during blastocyst implantation. • Endometrial cells were used in the comet assay to detect DNA damage. • CS2 exposure caused DNA damage and endometrial cell apoptosis. • DNA damage and endometrial cell apoptosis were responsible for embryo loss.

  3. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling, E-mail: shanglingwang@126.com

    2015-07-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H{sub 2}O{sub 2} production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling.

  4. [A first pilot study on the neonatal screening of primary immunodeficiencies in Spain: TRECS and KRECS identify severe T- and B-cell lymphopenia].

    Science.gov (United States)

    Olbrich, P; de Felipe, B; Delgado-Pecellin, C; Rodero, R; Rojas, P; Aguayo, J; Marquez, J; Casanovas, J; Sánchez, B; Lucena, J M; Ybot-Gonzalez, P; Borte, S; Neth, O

    2014-11-01

    Early diagnosis of primary immunodeficiency such as severe combined immunodeficiency (SCID) and X-linked agammaglobulinemia (XLA) improves outcome of affected infants/children. The measurement of T-cell receptor excision circles (TRECS) and kappa-deleting recombination excision circles (KRECS) can identify neonates with severe T or B-cell lymphopenia. To determine TRECS and KRECS levels from prospectively collected dried blood spot samples (DBS) and to correctly identify severe T and B-cell lymphopenia. Determination of TRECS and KRECS by multiplex PCR from neonates born in two tertiary hospitals in Seville between February 2014 and May 2014. PCR cut-off levels: TRECS1000 copies/μl. Internal (XLA, ataxia telangiectasia) and external (SCID) controls were included. A total of 1068 out of 1088 neonates (mean GA 39 weeks (38-40) and BW 3238g (2930-3520) were enrolled in the study. Mean (median, min/max) copies/μl, were as follows: TRECS 145 (132, 8/503), KRECS 82 (71, 7/381), and ACTB 2838 (2763, 284/7710). Twenty samples (1.87%) were insufficient. Resampling was needed in one neonate (0.09%), subsequently giving a normal result. When using lower cut-offs (TRECS<8 and KRECS<4 copies/μl), all the samples tested were normal and the internal and external controls were correctly identified. This is the first prospective pilot study in Spain using TRECS/KRECS/ACTB-assay, describing the experience and applicability of this method to identify severe lymphopenias. The ideal cut-off remains to be established in our population. Quality of sampling, storage and preparation need to be further improved. Copyright © 2014 Asociación Española de Pediatría. Published by Elsevier Espana. All rights reserved.

  5. Analysis of lysosomal membrane proteins exposed to melanin in HeLa cells

    OpenAIRE

    Bang, Seung Hyuck; Park, Dong Jun; Kim, Yang-Hoon; Min, Jiho

    2016-01-01

    Objectives There have been developed to use targeting ability for antimicrobial, anticancerous, gene therapy and cosmetics through analysis of various membrane proteins isolated from cell organelles. Methods It was examined about the lysosomal membrane protein extracted from lysosome isolated from HeLa cell treated by 100 ppm melanin for 24 hours in order to find associated with targeting ability to melanin using by 2-dimensional electrophoresis. Results The result showed 14 up-regulated (1.5...

  6. Influence of shear stress and size on viability of endothelial cells exposed to gold nanoparticles

    Science.gov (United States)

    Fede, C.; Albertin, Giovanna; Petrelli, L.; De Caro, R.; Fortunati, I.; Weber, V.; Ferrante, Camilla

    2017-09-01

    Screening nanoparticle toxicity directly on cell culture can be a fast and cheap technique. Nevertheless, to obtain results in accordance with those observed in live animals, the conditions in which cells are cultivated should resemble the one encountered in live systems. Microfluidic devices offer the possibility to satisfy this requirement, in particular with endothelial cell lines, because they are capable to reproduce the flowing media and shear stress experienced by these cell lines in vivo. In this work, we exploit a microfluidic device to observe how human umbilical vein endothelial cells (HUVEC) viability changes when subject to a continuous flow of culture medium, in which spherical citrate-stabilized gold nanoparticles of different sizes and at varying doses are investigated. For comparison, the same experiments are also run in multiwells where the cells do not experience the shear stress induced by the flowing medium. We discuss the results considering the influence of mode of exposure and nanoparticle size (24 and 13 nm). We observed that gold nanoparticles show a lower toxicity under flow conditions with respect to static and the HUVEC viability decreases as the nanoparticle surface area per unit volume increases, regardless of size.

  7. ATR controls cellular adaptation to hypoxia through positive regulation of hypoxia-inducible factor 1 (HIF-1) expression.

    Science.gov (United States)

    Fallone, F; Britton, S; Nieto, L; Salles, B; Muller, C

    2013-09-12

    Tumor cells adaptation to severe oxygen deprivation (hypoxia) plays a major role in tumor progression. The transcription factor HIF-1 (hypoxia-inducible factor 1), whose α-subunit is stabilized under hypoxic conditions is a key component of this process. Recent studies showed that two members of the phosphoinositide 3-kinase-related kinases (PIKKs) family, ATM (ataxia telangiectasia mutated) and DNA-PK (DNA-dependent protein kinase), regulate the hypoxic-dependent accumulation of HIF-1. These proteins initiate cellular stress responses when DNA damage occurs. In addition, it has been demonstrated that extreme hypoxia induces a replicative stress resulting in regions of single-stranded DNA at stalled replication forks and the activation of ATR (ataxia telangiectasia and Rad3 related protein), another member of the PIKKs family. Here, we show that even less severe hypoxia (0.1% O2) also induces activation of ATR through replicative stress. Importantly, in using either transiently silenced ATR cells, cells expressing an inactive form of ATR or cells exposed to an ATR inhibitor (CGK733), we demonstrate that hypoxic ATR activation positively regulates the key transcription factor HIF-1 independently of the checkpoint kinase Chk1. We show that ATR kinase activity regulates HIF-1α at the translational level and we find that the elements necessary for the regulation of HIF-1α translation are located within the coding region of HIF-1α mRNA. Finally, by using three independent cellular models, we clearly show that the loss of ATR expression and/or kinase activity results in the decrease of HIF-1 DNA binding under hypoxia and consequently affects protein expression levels of two HIF-1 target genes, GLUT-1 and CAIX. Taken together, our data show a new function for ATR in cellular adaptation to hypoxia through regulation of HIF-1α translation. Our work offers new prospect for cancer therapy using ATR inhibitors with the potential to decrease cellular adaptation in hypoxic

  8. Amiloride-sensitive sodium currents in fungiform taste cells of rats chronically exposed to nicotine.

    Science.gov (United States)

    Bigiani, A

    2015-01-22

    Many studies have demonstrated that chronic exposure to nicotine, one of the main components of tobacco smoke, has profound effects on the functionality of the mammalian taste system. However, the mechanisms underlying nicotine action are poorly understood. In particular no information is available on the chronic effect of nicotine on the functioning of taste cells, the peripheral detectors which transduce food chemicals into electrical signals to the brain. To address this issue, I studied the membrane properties of rat fungiform taste cells and evaluated the effect of long-term exposure to nicotine on the amiloride-sensitive sodium currents (ASSCs). These currents are mediated by the epithelial sodium channels (ENaC) thought to be important, at least in part, in the transduction of salty stimuli. Patch-clamp recording data indicated that ASSCs in taste cells from rats chronically treated with nicotine had a reduced amplitude compared to controls. The pharmacological and biophysical analysis of ASSCs revealed that amplitude reduction was not dependent on changes in amiloride sensitivity or channel ionic permeability, but likely derived from a decrease in the activity of ENaCs. Since these channels are considered to be sodium receptors in taste cells, my results suggest that chronic exposure to nicotine hampers the capability of these cells to respond to sodium ions. This might represent a possible cellular mechanism underlying the reduced taste sensitivity to salt typically found in smokers. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  9. System for exposing cultured cells to intermittent hypoxia utilizing gas permeable cultureware.

    Science.gov (United States)

    Polak, Jan; Studer-Rabeler, Karen; McHugh, Holly; Hussain, Mehboob A; Shimoda, Larissa A

    2015-07-01

    Tissue intermittent hypoxia (IH) occurs in obstructive sleep apnea, sickle cell anemia, physical exercise and other conditions. Poor gas solubility and slow diffusion through culture media hampers mimicking IH-induced transitions of O(2) in vitro. We aimed to develop a system enabling exposure of cultured cells to IH and to validate such exposure by real-time O(2) measurements and cellular responses. Standard 24-well culture plates and plates with bottoms made from a gas permeable film were placed in a heated cabinet. Desired cycling of O(2) levels was induced using programmable solenoids to purge mixtures of 95% N(2) + 5% CO(2) or 95% O(2) + 5% CO(2). Dissolved oxygen, gas pressure, temperature, and water evaporation were measured during cycling. IH-induced cellular effects were evaluated by hypoxia inducible factor (HIF) and NF-κB luciferase reporters in HEK296 cells and by insulin secretion in rat insulinoma cells. Oxygen cycling in the cabinet was translated into identical changes of O(2) at the well bottom in gas permeable, but not in standard cultureware. Twenty-four hours of IH exposure increased HIF (112%), NF-κB (111%) and insulin secretion (44%). Described system enables reproducible and prolonged IH exposure in cultured cells while controlling for important environmental factors.

  10. Sulindac enhances the killing of cancer cells exposed to oxidative stress.

    Directory of Open Access Journals (Sweden)

    Maria Marchetti

    2009-06-01

    Full Text Available Sulindac is an FDA-approved non-steroidal anti-inflammatory drug (NSAID that affects prostaglandin production by inhibiting cyclooxygenases (COX 1 and 2. Sulindac has also been of interest for more than decade as a chemopreventive for adenomatous colorectal polyps and colon cancer.Pretreatment of human colon and lung cancer cells with sulindac enhances killing by an oxidizing agent such as tert-butyl hydroperoxide (TBHP or hydrogen peroxide. This effect does not involve cyclooxygenase (COX inhibition. However, under the conditions used, there is a significant increase in reactive oxygen species (ROS within the cancer cells and a loss of mitochondrial membrane potential, suggesting that cell death is due to apoptosis, which was confirmed by Tunel assay. In contrast, this enhanced killing was not observed with normal lung or colon cells.These results indicate that normal and cancer cells handle oxidative stress in different ways and sulindac can enhance this difference. The combination of sulindac and an oxidizing agent could have therapeutic value.

  11. Induction of anchorage-independent growth in primary human cells exposed to protons or HZE ions separately or in dual exposures.

    Science.gov (United States)

    Sutherland, B M; Cuomo, N C; Bennett, P V

    2005-10-01

    Travelers on space missions will be exposed to a complex radiation environment that includes protons and heavy charged particles. Since protons are present at much higher levels than are heavy ions, the most likely scenario for cellular radiation exposure will be proton exposure followed by a hit by a heavy ion. Although the effects of individual ion species on human cells are being investigated extensively, little is known about the effects of exposure to both radiation types. One useful measure of mammalian cell damage is induction of the ability to grow in a semi-solid agar medium highly inhibitory to the growth of normal human cells, termed neoplastic transformation. Using primary human cells, we evaluated induction of soft-agar growth and survival of cells exposed to protons only or to heavy charged particles (600 MeV/nucleon silicon) only as well as of cells exposed to protons followed after a 4-day interval by silicon ions. Both ions alone efficiently transformed the human cells to anchorage-independent growth. Initial experiments indicate that the dose responses for neoplastic transformation of cells exposed to protons and then after 4 days to silicon ions appear similar to that of cells exposed to silicon ions alone.

  12. IFNγ/IL-10 co-producing cells dominate the CD4 response to malaria in highly exposed children.

    Directory of Open Access Journals (Sweden)

    Prasanna Jagannathan

    2014-01-01

    highly exposed children. These CD4(+ T cells may play important modulatory roles in the development of antimalarial immunity.

  13. Lack of Genomic Instability in Bone Marrow Cells of SCID Mice Exposed Whole-Body to Low-Dose Radiation

    Directory of Open Access Journals (Sweden)

    Elbert Whorton

    2013-04-01

    Full Text Available It is clear that high-dose radiation is harmful. However, despite extensive research, assessment of potential health-risks associated with exposure to low-dose radiation (at doses below or equal to 0.1 Gy is still challenging. Recently, we reported that 0.05 Gy of 137Cs gamma rays (the existing limit for radiation-exposure in the workplace was incapable of inducing significant in vivo genomic instability (measured by the presence of late-occurring chromosomal damage at 6 months post-irradiation in bone marrow (BM cells of two mouse strains, one with constitutively high and one with intermediate levels of the repair enzyme DNA-dependent protein-kinase catalytic-subunit (DNA-PKcs. In this study, we present evidence for a lack of genomic instability in BM cells of the severely combined-immunodeficiency (SCID/J mouse (which has an extremely low-level of DNA-PKcs activity exposed whole-body to low-dose radiation (0.05 Gy. Together with our previous report, the data indicate that low-dose radiation (0.05 Gy is incapable of inducing genomic instability in vivo (regardless of the levels of DNA-PKcs activity of the exposed mice, yet higher doses of radiation (0.1 and 1 Gy do induce genomic instability in mice with intermediate and extremely low-levels of DNA-PKcs activity (indicating an important role of DNA-PKcs in DNA repair.

  14. Damage Thresholds for Cultures RPE Cells Exposed to Lasers at 532 nm and 458 nm

    Science.gov (United States)

    2007-06-01

    in onhuman primate studies. Results of in vivo studies have hown that laser damage in the retina depends upon wave- ength, power level, and duration...condi- ions 37 °C; 95:5 air:CO2 using 1:1 DMEM/F12 media con- aining 10% fetal bovine serum, antibiotics, and 10-mM EPES buffer pH 7.4. Cells used...laser exposure similarly to RPE cells in nonhuman primate models. Our approach was to determine threshold ED50 radiant exposures for damage over a broad

  15. ATM Polymorphisms Predict Severe Radiation Pneumonitis in Patients With Non-Small Cell Lung Cancer Treated With Definitive Radiation Therapy

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Huihua [Department of Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan (China); Department of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Liao, Zhongxing, E-mail: zliao@mdanderson.org [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Liu, Zhensheng [Department of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Xu, Ting [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Wang, Qiming; Liu, Hongliang [Department of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Komaki, Ritsuko; Gomez, Daniel [Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Wang, Li-E [Department of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Wei, Qingyi, E-mail: qwei@mdanderson.org [Department of Epidemiology, The University of Texas MD Anderson Cancer Center, Houston, Texas (United States)

    2013-03-15

    Purpose: The ataxia telangiectasia mutated (ATM) gene mediates detection and repair of DNA damage. We investigated associations between ATM polymorphisms and severe radiation-induced pneumonitis (RP). Methods and Materials: We genotyped 3 potentially functional single nucleotide polymorphisms (SNPs) of ATM (rs1801516 [D1853N/5557G>A], rs189037 [-111G>A] and rs228590) in 362 patients with non-small cell lung cancer (NSCLC), who received definitive (chemo)radiation therapy. The cumulative severe RP probabilities by genotypes were evaluated using the Kaplan-Meier analysis. The associations between severe RP risk and genotypes were assessed by both logistic regression analysis and Cox proportional hazard model with time to event considered. Results: Of 362 patients (72.4% of non-Hispanic whites), 56 (15.5%) experienced grade ≥3 RP. Patients carrying ATM rs189037 AG/GG or rs228590 TT/CT genotypes or rs189037G/rs228590T/rs1801516G (G-T-G) haplotype had a lower risk of severe RP (rs189037: GG/AG vs AA, adjusted hazard ratio [HR] = 0.49, 95% confidence interval [CI], 0.29-0.83, P=.009; rs228590: TT/CT vs CC, HR=0.57, 95% CI, 0.33-0.97, P=.036; haplotype: G-T-G vs A-C-G, HR=0.52, 95% CI, 0.35-0.79, P=.002). Such positive findings remained in non-Hispanic whites. Conclusions: ATM polymorphisms may serve as biomarkers for susceptibility to severe RP in non-Hispanic whites. Large prospective studies are required to confirm our findings.

  16. DJ1 Expression Downregulates in Neuroblastoma Cells (SK-N-MC Chronically Exposed to HIV-1 and Cocaine.

    Directory of Open Access Journals (Sweden)

    Upal eRoy

    2015-07-01

    Full Text Available Background: HIV-associated neurological disorder (HAND has long been recognized as a consequence of Human Immunodeficiency Virus (HIV infection in the brain. The pathology of HAND gets more complicated with the recreational drug use such as cocaine. Recent studies have suggested multiple genetic influences involved in the pathology of addiction and HAND but only a fraction of the entire genetic risk has been investigated so far. In this regard, role of DJ1 protein (a gene linked to autosomal recessive early-onset Parkinson’s disease in regulating dopamine transmission and reactive oxygen species (ROS production in neuronal cells will be worth investigating in HIV-1 and cocaine exposed microenvironment. Being a very abundant protein in the brain, DJ1 could serve as a potential marker for early detection of HIV-1 and/or cocaine related neurological disorder.Methods: In vitro analysis was done to observe the effect of HIV-1 and/or cocaine on DJ1 protein expression in neuroblastoma cells (SK-N-MC. Gene expression and protein analysis of DJ1 was done on the HIV infected and/or cocaine treated SK-N-MC and compared to untreated cells using real time PCR, Western Blot and flow cytometry.Results: Gene expression and protein analysis indicated that there was a significant decrease in DJ1 expression in SK-N-MC chronically exposed to HIV-1 and/or cocaine.Conclusion: This is the first study to establish that DJ1 expression level in the neuronal cells significantly decreased in presence of HIV-1and/or cocaine indicating oxidative stress level of dopamine neurons.

  17. Toxic cyanobacterial cells containing microcystins induce oxidative stress in exposed tilapia fish (Oreochromis sp.) under laboratory conditions.

    Science.gov (United States)

    Jos, Angeles; Pichardo, Silvia; Prieto, Ana I; Repetto, Guillermo; Vázquez, Carmen M; Moreno, Isabel; Cameán, Ana M

    2005-04-30

    The effects of microcystins from cyanobacterial cells on various oxidative stress biomarkers in liver, kidney and gill tissues in freshwater tilapia fish (Oreochromis sp.) were investigated under laboratory conditions. Microcystins are a family of cyclic peptide toxins produced by species of freshwater cyanobacteria (blue-green algae). Fish were exposed to the cyanobacterial cells in two ways: mixed with a commercial fish food or crushed into a commercial fish food so that the toxins were released. Two different exposure times were studied: 14 and 21 days. The oxidative status of fish was evaluated by analyzing the level of lipid peroxidation (LPO), as well as the activities of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR). The findings of the present investigation show that microcystins induce oxidative stress in a time-dependent manner and that the type of administration of the cyanobacterial cells influences the extent of these effects. Thus, the crushed cyanobacterial cells (released toxins) induced the antioxidant defences studied and increased the LPO level to a greater extent than the non-crushed cells. The liver was the most affected organ followed by kidney and gills. These results together with reports that fish can accumulate microcystins mean that cyanobacterial blooms are an important health, environmental and economic problem.

  18. Gypenosides Protected the Neural Stem Cells in the Subventricular Zone of Neonatal Rats that Were Prenatally Exposed to Ethanol

    Directory of Open Access Journals (Sweden)

    Lun Dong

    2014-11-01

    Full Text Available Fetal alcohol spectrum disorder (FASD can cause severe mental retardation in children who are prenatally exposed to ethanol. The effects of prenatal and early postnatal ethanol exposure on adult hippocampal neurogenesis have been investigated; however, the effects of prenatal ethanol exposure on the subventricular zone (SVZ have not. Gypenosides (GPs have been reported to have neuroprotective effects in addition to other bioactivities. The effects of GPs on neural stem cells (NSCs in the FASD model are unknown. Here, we test the effect of prenatal ethanol exposure on the neonatal SVZ, and the protection potential of GPs on NSCs in FASD rats. Our results show that prenatal ethanol exposure can suppress the cell proliferation and differentiation of neural stem cells in the neonatal SVZ and that GPs (400 mg/kg/day can significantly increase the cell proliferation and differentiation of neural stem cells inhibited by ethanol. Our data indicate that GPs have neuroprotective effects on the NSCs and can enhance the neurogenesis inhibited by ethanol within the SVZ of neonatal rats. These findings provide new evidence for a potential therapy involving GPs for the treatment of FASD.

  19. CC-115, a dual inhibitor of mTOR kinase and DNA-PK, blocks DNA damage repair pathways and selectively inhibits ATM-deficient cell growth in vitro.

    Science.gov (United States)

    Tsuji, Toshiya; Sapinoso, Lisa M; Tran, Tam; Gaffney, Bonny; Wong, Lilly; Sankar, Sabita; Raymon, Heather K; Mortensen, Deborah S; Xu, Shuichan

    2017-09-26

    CC-115, a selective dual inhibitor of the mammalian target of rapamycin (mTOR) kinase and DNA-dependent protein kinase (DNA-PK), is undergoing Phase 1 clinical studies. Here we report the characterization of DNA-PK inhibitory activity of CC-115 in cancer cell lines. CC-115 inhibits auto-phosphorylation of the catalytic subunit of DNA-PK (DNA-PKcs) at the S2056 site (pDNA-PK S2056), leading to blockade of DNA-PK-mediated non-homologous end joining (NHEJ). CC-115 also indirectly reduces the phosphorylation of ataxia-telangiectasia mutated kinase (ATM) at S1981 and its substrates as well as homologous recombination (HR). The mTOR kinase and DNA-PK inhibitory activity of CC-115 leads to not only potent anti-tumor activity against a large panel of hematopoietic and solid cancer cell lines but also strong induction of apoptosis in a subset of cancer lines. Mechanistically, CC-115 prevents NHEJ by inhibiting the dissociation of DNA-PKcs, X-ray repair cross-complementing protein 4 (XRCC4), and DNA ligase IV from DNA ends. CC-115 inhibits colony formation of ATM-deficient cells more potently than ATM-proficient cells, indicating that inhibition of DNA-PK is synthetically lethal with the loss of functional ATM. In conclusion, CC-115 inhibits both mTOR signaling and NHEJ and HR by direct inhibition of DNA-PK. The mechanistic data not only provide selection of potential pharmacodynamic (PD) markers but also support CC-115 clinical development in patients with ATM-deficient tumors.

  20. Uroporphyrinogen-I-synthetase activity in red blood cells of lead-exposed workers

    Energy Technology Data Exchange (ETDEWEB)

    El-Waseef, A.

    1982-01-01

    Lead-exposed (n . 26) and control (n . 12) subjects were investigated for their blood lead concentration erythrocyte 5-amino-laevulinic acid dehydratase (5-ALAD) and erythrocyte uroporphyrinogen-I-synthetase (URO-I-S) activity; 5-amino-laevulinic acid (5-ALA) and porphobilinogen (PBG) were used as substrates in the synthetase assay. In the lead workers erythrocyte 5-ALA dehydratase was grossly inhibited but with PBG as substrate the synthetase activity was not significantly different from the control group. With 5-ALA as substrate the synthetase assay showed marked inhibition. Addition of zinc (0.1 mmol/l) and dithiotheritol (0.5 mmol/l) brought the activities of both the dehydratase and synthetase (using 5-ALA as substrate) back into the ranges seen in the control group. With porphobilinogen as substrate higher concentrations of zinc caused inhibition of the synthetase, whilst reduction of added zinc to 0.01 mmol/l resulted in stimulation of the synthetase. A good correlation (r . 0.87) was obtained in synthetase assay when PBG and 5-aminolaevulinate (with added zinc and dithiothreitol) were used as substrates. With these additions 5-ALA may be used as a substrate in the URO-I-S assay in the investigation of latent cases of acute intermittent porphyria.

  1. In vitro measurements of oxygen consumption rates in hTERT-RPE cells exposed to low levels of red light

    Science.gov (United States)

    Wigle, Jeffrey C.; Castellanos, Cherry C.

    2016-03-01

    Exposure to 2.88 J/cm2 of red light induces an adaptive response against a lethal pulse of 2.0 μm laser radiation in hTERT-RPE cells in vitro, but not in a knockdown mutant for vascular endothelial growth factor c (VEGF-C). The generally accepted initiation sequence for photobiomodulation is that absorption of red light by cytochome c oxidase (CCOX) of the electron transport chain increases the binding affinity of CCOX for O2 vs. nitric oxide (NO). This results in displacement of NO by O2 in the active site of CCOX, thereby increasing cellular respiration and intracellular ATP. We've previously reported that red-light exposure induces a small, but consistently reproducible, increase in NO levels in these cells. But the relative importance of NO and oxidative phosphorylation is unclear because little is known about the relative contributions of NO and ATP to the response. However, if NO dissociation from CCOX actually increases oxidative phosphorylation, one should see a corresponding increase in oxygen consumption. A Seahorse Extracellular Flux Analyzer was used to measure oxygen consumption rates (OCR) in normal and mutant cells as a proxy for oxidative phosphorylation. Both basal respiration and maximum respiration rates in normal cells are significantly higher than in the mutant. The normal cells have a significant amount of "excess capacity," whereas the VEGF-C(KD) have little or none. The OCR in exposed normal cells is lower than in unexposed cells when measured immediately after exposure. The exposures used for these experiments had no effect on the OCR in mutant cells.

  2. Biomonitoring of genotoxic and cytotoxic effects of gingival epithelial cells exposed to digital panoramic radiography

    Directory of Open Access Journals (Sweden)

    Anuradha Pai

    2012-01-01

    Full Text Available Objective: The aim of this study was to evaluate genotoxic and cytotoxic effects of low level ionizing radiation used in digital panoramic radiography on gingival epithelial cells. Materials and Methods: We included 50 healthy individuals advised for digital panoramic radiography for diagnostic purpose were included in this study. Demographic data and personal history of all subjects were recorded in a proforma before the examination. Gingival epithelial cells were obtained by gentle scraping with a modified cytobrush immediately before X-ray exposure and 10 ± 2 days later. Cytological preparations were stained according to the Feulgen/fast green method and analyzed under a light microscope. Micronuclei and degenerative nuclear alterations (pyknosis, karyolysis, karyorrhexis and condensed chromatin were scored. Results: The frequency of formation of micronuclei was not significant with regard to age, gender and after exposure to digital panoramic radiography ( P = 0.276. However this study showed significant increase in the frequencies of nuclear alterations like karyorrhexis, pyknosis, condensed chromatin, karyolysis and indicative of cell death ( P < 0.001. Conclusion: Panoramic radiographic examination does not induce genotoxic effect like micronuclei, but it does induce cytotoxic effects leading to cell death.

  3. c-jun gene expression in human cells exposed to either ionizing radiation or hydrogen peroxide

    Energy Technology Data Exchange (ETDEWEB)

    Collart, F.R.; Horio, M.; Huberman, E.

    1993-06-01

    We investigated the role of reactive oxygen intermediates (ROIs) and protein kinase C (PKC) in radiation- and H{sub 2}O{sub 2}-evoked c-jun gene expression in human HL-205 cells. This induction of c-jun gene expression could be prevented by pretreatment of the cells with Nacetylcysteine (an antioxidant) or H7 (a PKC and PKA inhibitor) but not by HA1004, a PKA inhibitor, suggesting a role for ROls and PKC in mediating c-jun gene expression. We also investigated potential differences in c-jun gene expression in a panel of normal and tumor cells untreated or treated with ionizing radiation or H{sub 2}O{sub 2}. Treatment with radiation or H{sub 2}O{sub 2} produced a varied response, from some reduction to an increase of more than an order of magnitude in the steady-state level of c-jun mRNA. These data indicate that although induction of c-jun may be a common response to ionizing radiation and H{sub 2}O{sub 2}, this response was reduced or absent in some cell types.

  4. Disruption of glucagon receptor signaling causes hyperaminoacidemia exposing a possible liver - alpha-cell axis

    DEFF Research Database (Denmark)

    Galsgaard, Katrine D; Winther-Sørensen, Marie; Ørskov, Cathrine

    2018-01-01

    Glucagon secreted from the pancreatic alpha-cells is essential for regulation of blood glucose levels. However, glucagon may play an equally important role in the regulation of amino acid metabolism by promoting ureagenesis. We hypothesized that disruption of glucagon receptor signaling would lea...

  5. Protective effect of a mexican propolis on MDBK cells exposed to ...

    African Journals Online (AJOL)

    Background: In this paper, the evaluation of the antiviral effect of a Mexican propolis on Pseudo Rabies Virus (PRV) was performed by infecting cell cultures of MDBK. Materials and Methods: First, the level of cytotoxicity of the ethanol extract of propolis (EEP) was determined, subsequently, infective dose of PRV strain ...

  6. The morphological and molecular changes of brain cells exposed to direct current electric field stimulation.

    Science.gov (United States)

    Pelletier, Simon J; Lagacé, Marie; St-Amour, Isabelle; Arsenault, Dany; Cisbani, Giulia; Chabrat, Audrey; Fecteau, Shirley; Lévesque, Martin; Cicchetti, Francesca

    2014-12-07

    The application of low-intensity direct current electric fields has been experimentally used in the clinic to treat a number of brain disorders, predominantly using transcranial direct current stimulation approaches. However, the cellular and molecular changes induced by such treatment remain largely unknown. Here, we tested various intensities of direct current electric fields (0, 25, 50, and 100V/m) in a well-controlled in vitro environment in order to investigate the responses of neurons, microglia, and astrocytes to this type of stimulation. This included morphological assessments of the cells, viability, as well as shape and fiber outgrowth relative to the orientation of the direct current electric field. We also undertook enzyme-linked immunosorbent assays and western immunoblotting to identify which molecular pathways were affected by direct current electric fields. In response to direct current electric field, neurons developed an elongated cell body shape with neurite outgrowth that was associated with a significant increase in growth associated protein-43. Fetal midbrain dopaminergic explants grown in a collagen gel matrix also showed a reorientation of their neurites towards the cathode. BV2 microglial cells adopted distinct morphological changes with an increase in cyclooxygenase-2 expression, but these were dependent on whether they had already been activated with lipopolysaccharide. Finally, astrocytes displayed elongated cell bodies with cellular filopodia that were oriented perpendicularly to the direct current electric field. We show that cells of the central nervous system can respond to direct current electric fields both in terms of their morphological shape and molecular expression of certain proteins, and this in turn can help us to begin understand the mechanisms underlying the clinical benefits of direct current electric field. © The Author 2015. Published by Oxford University Press on behalf of CINP.

  7. Intracellular ice and cell survival in cryo-exposed embryonic axes of recalcitrant seeds of Acer saccharinum: an ultrastructural study of factors affecting cell and ice structures

    CSIR Research Space (South Africa)

    Wesley-Smith, J

    2014-03-01

    Full Text Available Annals of Botany 113: 695–709, 2014 doi:10.1093/aob/mct284, available online at www.aob.oxfordjournals.org Intracellular ice and cell survival in cryo-exposed embryonic axes of recalcitrant seeds of Acer saccharinum: an ultrastructural study... of factors affecting cell and ice structures JamesWesley-Smith1,2, Patricia Berjak1, N.W. Pammenter1 and ChristinaWalters3,* 1Plant Germplasm Conservation Research, School of Life Sciences, University of KwaZulu- Natal(Westville Campus), Durban, 4001...

  8. Induction of Cell Death through Alteration of Oxidants and Antioxidants in Epithelial Cells Exposed to High Energy Protons

    Science.gov (United States)

    Ramesh, Govindarajan; Wu, Honglu

    2012-01-01

    Radiation affects several cellular and molecular processes including double strand breakage, modifications of sugar moieties and bases. In outer space, protons are the primary radiation source which poses a range of potential health risks to astronauts. On the other hand, the use of proton radiation for tumor radiation therapy is increasing as it largely spares healthy tissues while killing tumor tissues. Although radiation related research has been conducted extensively, the molecular toxicology and cellular mechanisms affected by proton radiation remain poorly understood. Therefore, in the present study, we irradiated rat epithelial cells (LE) with different doses of protons and investigated their effects on cell proliferation and cell death. Our data showed an inhibition of cell proliferation in proton irradiated cells with a significant dose dependent activation and repression of reactive oxygen species (ROS) and antioxidants, glutathione and superoxide dismutase respectively as compared to control cells. In addition, apoptotic related genes such as caspase-3 and -8 activities were induced in a dose dependent manner with corresponding increased levels of DNA fragmentation in proton irradiated cells than control cells. Together, our results show that proton radiation alters oxidant and antioxidant levels in the cells to activate apoptotic pathway for cell death.

  9. [Ultrastructural study of the interstitial cells of the internal kidney medulla exposed to indomethacin].

    Science.gov (United States)

    Anavi, B; Dragiev, M

    1981-01-01

    Unusual cylindrical bodies-megatubules, which get in touch with lipid granules with granular endoplasmatic reticulum among themselves and with the pores of the nuclear and cellular membrane, are described in the interstitial cells of the kidneys of the control rats. After a suppression of prostaglandin synthesis with indomethacin (5 mg/kg venously of intraperitonealy per day for 6 days) there was statistically significant increase of the mass of the compact lipids in the interstitial cells (hypergranulation as the volume of the granule was preserved) - a proof that the granules contained precursors of renal prostaglandins. The nuclei were presented with round contours, while the perinuclear spaces were narrowed. The Goldgi apparatus and granular endoplasmic reticulum hyperthrophied. The megatubules atrophied. The authors suggest that they have prostaglandin synthetase activity.

  10. Cultured bovine aortic endothelial cells show increased histamine metabolism when exposed to oscillatory shear stress.

    Science.gov (United States)

    Skarlatos, S I; Hollis, T M

    1987-03-01

    Oscillatory shear stress applied to the lining of blood vessels causes endothelial cell injury, one of the essential postulated prerequisites to the development of atherosclerosis. The purpose of this investigation was to study effects of shear stress on bovine aortic endothelial cells (BAEC), in vitro, for varying lengths of time (6 h, 12 h, 24 h) on BAEC histamine content (HC) and histidine decarboxylase activity (HD). Low intensity stress (1.6 dynes/cm2) as well as intermediate and high intensity shear stresses (3.5 dynes/cm2 and 7.6 dynes/cm2) resulted in an accelerated HD (281%) and elevated HC (144%). These data indicate that oscillatory shear stress produces increases in histamine metabolism.

  11. DNA Repair in Human Cells Exposed to Combinations of Carcinogenic Agents

    Energy Technology Data Exchange (ETDEWEB)

    Setlow, R. B.; Ahmed, F. E.

    1980-01-01

    Normal human and XP2 fibroblasts were treated with UV plus UV-mimetic chemicals. The UV dose used was sufficient to saturate the UV excision repair system. Excision repair after combined treatments was estimated by unscheduled DNA synthesis, BrdUrd photolysis, and the loss of sites sensitive to a UV specific endonuclease. Since the repair of damage from UV and its mimetics is coordinately controlled we expected that there would be similar rate-limiting steps in the repair of UV and chemical damage and that after a combined treatment the total amount of repair would be the same as from UV or the chemicals separately. The expectation was not fulfilled. In normal cells repair after a combined treatment was additive whereas in XP cells repair after a combined treatment was usually less than after either agent separately. The chemicals tested were AAAF, DMBA-epoxide, 4NQO, and ICR-170.

  12. Modeling of Nanoparticle-Mediated Electric Field Enhancement Inside Biological Cells Exposed to AC Electric Fields

    Science.gov (United States)

    Tiwari, Pawan K.; Kang, Sung Kil; Kim, Gon Jun; Choi, Jun; Mohamed, A.-A. H.; Lee, Jae Koo

    2009-08-01

    We present in this article the effect of alternating electric field at kilohertz (kHz) and megahertz (MHz) frequencies on the biological cells in presence and absence of nanoparticles. The induced electric field strength distribution in the region around cell membrane and nucleus envelope display different behavior at kHz and MHz frequencies. The attachment of gold nanoparticles (GNPs), especially gold nanowires around the surface of nucleus induce enhanced electric field strengths. The induced field strengths are dependent on the length of nanowire and create varying field regions when the length of nanowire is increased from 2 to 4 µm. The varying nanowire length increased the induced field strengths inside nucleoplasm and region adjacent to the nucleus in the cytoplasm. We investigated a process of electrostatic disruption of nucleus membrane when the induced electric field strength across the nucleus exceeds its tensile strength.

  13. Modeling of a single red blood cell thermal reaction exposed to infrared laser tweezers

    Science.gov (United States)

    Seteikin, A.; Krasnikov, I.; Bernhardt, I.

    2013-02-01

    Continuous-wave laser micro-beams are generally used as diagnostic tools in laser scanning microscopes or in the case of near-infrared (NIR) micro-beams, as optical traps for cell manipulation and force characterization. Because single beam traps are created with objectives of high numerical aperture, typical trapping intensities and photon flux densities are in the order of 106 W/cm2 and 103 cm-2s-1, respectively. The main idea of our theoretical study was to investigate the thermal reaction of RBCs irradiated by laser micro-beam. The study is supported by the fact that many experiments have been carried out with RBCs in laser NIR tweezers. In the present work it has been identified that the laser affects a RBC with a density of absorbed energy at approximately 107 J/cm3, which causes a temperature rise in the cell of about 7 - 12 °C.

  14. CYTOKINESIS-BLOCK MICRONUCLEUS ASSAY IN HUMAN GLIOMA CELLS EXPOSED TO RADIATION

    Directory of Open Access Journals (Sweden)

    Jerzy Slowinski

    2011-05-01

    Full Text Available Biological tests are efficient in reflecting the biological influences of several types of generally harmful exposures. The micronucleus assay is widely used in genotoxicity studies or studies on genomic damage in general. We present methodological aspects of cytokinesis-block micronucleus assay performed in human gliomas irradiated in vitro. Eight human glioblastoma cell lines obtained from DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Germany were gamma-irradiated (60Co over a dose range of 0-10 Gy. Cytokinesis-block micronucleus assay was performed to quantitate cytogenetic damage. The cells were fixed directly on dishes, stained with fluorochrome DAPI and evaluated under fluorescent and phase contrast microscope. The micronucleus frequency was expressed as a micronuclei (MN per binucleated cell (BNC ratio, calculated after scoring at least 100 BNC per dish. The frequency of spontaneous MN ranged from 0.17 to 0.613 (mean: 0.29 ± 0.14. After irradiation increase of MN frequency in the range of 0.312 - 2.241 (mean: 0.98 ± 0.68 was found at 10 Gy. Gliomas are extremely heterogenous in regard to cytogenetic effects of irradiation, as shown in this study by cytokinesis-block micronucleus assay. This test is easily performed on irradiated glioma cell lines and can assist in determining their radiosensitivity. However, in order to obtain reliable and reproducible results, precise criteria for MN scoring must be strictly followed. Simultaneous use of fluorescent and phase contrast equipment improves imaging of morphological details and can further optimize MN scoring.

  15. Effect of roflumilast on inflammatory cells in the lungs of cigarette smoke-exposed mice

    OpenAIRE

    Martorana, Piero A; Lunghi, Benedetta; Lucattelli, Monica; De Cunto, Giovanna; Beume, Rolf; Lungarella, Giuseppe

    2008-01-01

    Abstract Background We reported that roflumilast, a phosphodiesterase 4 inhibitor, given orally at 5 mg/kg to mice prevented the development of emphysema in a chronic model of cigarette smoke exposure, while at 1 mg/kg was ineffective. Here we investigated the effects of roflumilast on the volume density (VV) of the inflammatory cells present in the lungs after chronic cigarette smoke exposure. Methods Slides were obtained from blocks of the previous study and VV was assessed immunohistochemi...

  16. Extracellular signal-regulated kinases 1 and 2 activation in endothelial cells exposed to cyclic strain

    Science.gov (United States)

    Ikeda, M.; Takei, T.; Mills, I.; Kito, H.; Sumpio, B. E.

    1999-01-01

    The aim of this study was to determine whether extracellular signal-regulated kinases 1/2 (ERK1/ERK2) are activated and might play a role in enhanced proliferation and morphological change induced by strain. Bovine aortic endothelial cells (BAEC) were subjected to an average of 6 or 10% strain at a rate of 60 cycles/min for up to 4 h. Cyclic strain caused strain- and time-dependent phosphorylation and activation of ERK1/ERK2. Peak phosphorylation and activation of ERK1/ERK2 induced by 10% strain were at 10 min. A specific ERK1/ERK2 kinase inhibitor, PD-98059, inhibited phosphorylation and activation of ERK1/ERK2 but did not inhibit the increased cell proliferation and cell alignment induced by strain. Treatment of BAEC with 2,5-di-tert-butyl-1, 4-benzohydroquinone, to deplete inositol trisphosphate-sensitive calcium storage, and gadolinium chloride, a Ca2+ channel blocker, did not inhibit the activation of ERK1/ERK2. Strain-induced ERK1/ERK2 activation was partly inhibited by the protein kinase C inhibitor calphostin C and completely inhibited by the tyrosine kinase inhibitor genistein. These data suggest that 1) ERK1/ERK2 are not critically involved in the strain-induced cell proliferation and orientation, 2) strain-dependent activation of ERK1/ERK2 is independent of intracellular and extracellular calcium mobilization, and 3) protein kinase C activation and tyrosine kinase regulate strain-induced activation of ERK1/ERK2.

  17. Proteomic Analysis of MCF-7 Breast Cancer Cell Line Exposed To Leptin

    Directory of Open Access Journals (Sweden)

    A. Valle

    2011-01-01

    Full Text Available Background: Obesity is a well-known factor risk for breast cancer in postmenopausal women. Circulating leptin levels are increased in obese and it has been suggested to play an important role in mammary tumor formation and progression. To contribute to the understanding of the molecular mechanisms underlying leptin action in breast cancer, our aim was to identify proteins regulated by leptin in MCF-7 human breast cancer cells. Methods: We used two-dimensional gel electrophoresis (2-DE and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS to identify proteins affected by leptin. Results: Thirty proteins were found differentially expressed in MCF-7 cells after 48 h leptin exposure. Proteins regulated by leptin included proteins previously implicated in breast cancer such as catechol-o-methyltransferase, cathepsin D, hsp27, serine/threonine-protein phosphatase and regulatory proteins of the Ras signaling pathway. Proteins involved in other cellular functions such as stress response, cytosqueleton remodeling and proteins belonging to ubiquitin-proteasome system, were also identified. Furthermore, leptin-treated cells showed a substantial uptake of the serum carrier proteins albumin and alpha-2-HS-glycoprotein. Conclusions: This screening reveals that leptin influences the levels of key proteins involved in breast cancer which opens new avenues for the study of the molecular mechanisms linking obesity to breast cancer.

  18. Photocatalytic Oxidation of Triiodide in UVA-Exposed Dye-Sensitized Solar Cells

    Directory of Open Access Journals (Sweden)

    Matthew Carnie

    2012-01-01

    Full Text Available UVA irradiation of glass mounted dye-sensitized solar cells without UV filtration causes failure within 400 hours of light exposure. The failure mode is shown to relate to consumption of I3−, which is directly related to TiO2 photo-catalysis. The onset of failure is easily determined from electrochemical impedance data where the recombination resistance of the TiO2/electrolyte back reaction drops markedly prior to the onset of degradation. At the point of complete cell failure this impedance value then dramatically increases as there is no longer an interfacial reaction possible between the TiO2 and the I3− depleted electrolyte. Device failure is most rapid for cells under electrical load indicating that the degradation of the electrolyte is related to photogenerated hole production by excitation of the TiO2. Once depleted by UV exposure, the I3− can be regenerated by simple application of a reverse bias which can restore severely UV degraded devices to near original working conditions.

  19. Protective Pleiotropic Effect of Flavonoids on NAD+ Levels in Endothelial Cells Exposed to High Glucose

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    Daniëlle M. P. H. J. Boesten

    2015-01-01

    Full Text Available NAD+ is important for oxidative metabolism by serving as an electron transporter. Hyperglycemia decreases NAD+ levels by activation of the polyol pathway and by overactivation of poly(ADP-ribose-polymerase (PARP. We examined the protective role of three structurally related flavonoids (rutin, quercetin, and flavone during high glucose conditions in an in vitro model using human umbilical vein endothelial cells (HUVECs. Additionally we assessed the ability of these flavonoids to inhibit aldose reductase enzyme activity. We have previously shown that flavonoids can inhibit PARP activation. Extending these studies, we here provide evidence that flavonoids are also able to protect endothelial cells against a high glucose induced decrease in NAD+. In addition, we established that flavonoids are able to inhibit aldose reductase, the key enzyme in the polyol pathway. We conclude that this protective effect of flavonoids on NAD+ levels is a combination of the flavonoids ability to inhibit both PARP activation and aldose reductase enzyme activity. This study shows that flavonoids, by a combination of effects, maintain the redox state of the cell during hyperglycemia. This mode of action enables flavonoids to ameliorate diabetic complications.

  20. Cytogenetic biomonitoring of oral mucosa cells from adults exposed to dental X-rays.

    Science.gov (United States)

    Ribeiro, Daniel A; Angelieri, Fernanda

    2008-07-01

    Although it has been clearly demonstrated that X-rays play a key role in diagnosing medical and dental problems, this type of ionizing radiation is also able to induce noxious activities, such as genetic damage. The aim of the present study was to evaluate DNA damage (micronucleus) and cellular death in exfoliated buccal mucosa cells from healthy individuals (smokers and nonsmokers) following dental X-ray exposure. A total of 39 healthy people who had submitted to panoramic dental radiography were included in the study: 9 smokers and 30 nonsmokers. The results indicated no significant statistically differences (P>0.05) in micronucleated oral mucosa cells before and after dental X-ray exposure. On the other hand, X-ray exposure did increase other nuclear alterations closely related to cytotoxicity, such as karyorrhexis, pyknosis, and karyolysis. It seems that cigarette smoke did not affect X-ray outcomes induced in buccal cells. These data indicate that dental panoramic radiography may not induce chromosomal damage, but it is able to promote cytotoxicity. Because cellular death is considered a prime mechanism in nongenotoxic mechanisms of carcinogenesis, dental X-ray should be used only when necessary.

  1. [Protective effects of amygdalin on hyperoxia-exposed type II alveolar epithelial cells isolated from premature rat lungs in vitro].

    Science.gov (United States)

    Chang, Li-wen; Zhu, Hua-ping; Li, Wen-bin; Liu, Han-chu; Zhang, Qian-shen; Chen, Hong-bing

    2005-02-01

    To analyze the effect of hyperoxia on the proliferation and surfactant associated protein messenger RNA levels of type II alveolar epithelial cells (AECIIs) of premature rat, and to investigate the effect of amygdalin on the change resulted from hyperoxia in AECIIs isolated from premature rat lung in vitro. The lung tissue of 20-day fetal rat was digested by trypsin and collagenase. AECIIs and lung fibroblasts (LFs) were isolated and purified at different centrifugal force and different adherence, then cultured. The nature of the cultures was identified by cytokeratin staining, vimentin staining and transmission electron micrography. For establishing hyperoxia-exposed cell model, purified AECIIs were cultured for 24 hours after culture flasks were filled with 95% oxygen-5% CO2 at 3 L/min for 10 min, and then sealed. Oxygen concentrations were tested in CYS-1 digital oxygen monitor after 24 hours of exposure. A sample was discarded if its oxygen concentration was amygdalin at various concentrations. DNA content, protein expression of proliferating cell nuclear antigen (PCNA) and mRNA levels of SPs of AECIIs were analyzed with flow cytometric assay, Western blot and reverse transcription polymerase chain reaction (RT-PCR) respectively after 24 hours of air or hyperoxia exposure in the presence or absence of 200 micromol/L amygdalin. Excellent yields of highly purified, culturable AECIIs could be obtained from 20-day fetal lungs. The expression of cytokeratin in AECIIs was positive and that of vimentin negative by immunocytochemistry. Those, however, in LFs were just opposite. Lamellar bodies in purified AECIIs were revealed by transmission electron micrography. The established hyperoxia-exposed cell model assured the oxygen concentrations of culture flasks more than 90%. Amygdalin at the concentration range from 50 micromol/L to 200 micromol/L stimulated the proliferation of AECIIs in a dose-dependent manner; however, at the concentration of 400 micromol/L inhibited

  2. Bioinformatic Analysis of Differential Protein Expression in Calu-3 Cells Exposed to Carbon Nanotubes

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    Pin Li

    2013-10-01

    Full Text Available Carbon nanomaterials are widely produced and used in industry, medicine and scientific research. To examine the impact of exposure to nanoparticles on human health, the human airway epithelial cell line, Calu-3, was used to evaluate changes in the cellular proteome that could account for alterations in cellular function of airway epithelia after 24 hexposure to 10 μg/mL and 100 ng/mLof two common carbon nanoparticles, single- and multi-wall carbon nanotubes (SWCNT, MWCNT. After exposure to the nanoparticles, label-free quantitative mass spectrometry (LFQMS was used to study the differential protein expression. Ingenuity Pathway Analysis (IPA was used to conduct a bioinformaticanalysis of proteins identified in LFQMS. Interestingly, after exposure to ahigh concentration (10 mg/mL; 0.4 mg/cm2 of MWCNT or SWCNT, only 8 and 13 proteins, respectively, exhibited changes in abundance. In contrast, the abundance of hundreds of proteins was altered in response to a low concentration (100 ng/mL; 4 ng/cm2 of either CNT. Of the 281 and 282 proteins that were significantly altered in response to MWCNT or SWCNT respectively, 231 proteins were the same. Bioinformatic analyses found that the proteins in common to both nanotubes occurred within the cellular functions of cell death and survival, cell-to-cell signaling and interaction, cellular assembly and organization, cellular growth and proliferation, infectious disease, molecular transport and protein synthesis. The majority of the protein changes represent a decrease in amount suggesting a general stress response to protect cells. The STRING database was used to analyze the various functional protein networks. Interestingly, some proteins like cadherin 1 (CDH1, signal transducer and activator of transcription 1 (STAT1, junction plakoglobin (JUP, and apoptosis-associated speck-like protein containing a CARD (PYCARD, appear in several functional categories and tend to be in the center of the networks. This

  3. Neuroprotective effects of melatonin and omega-3 on hippocampal cells prenatally exposed to 900 MHz electromagnetic fields.

    Science.gov (United States)

    Erdem Koç, Gülüna; Kaplan, Suleyman; Altun, Gamze; Gümüş, Hasan; Gülsüm Deniz, Ömür; Aydin, Isinsu; Emin Onger, Mehmet; Altunkaynak, Zuhal

    2016-10-01

    Adverse effects on human health caused by electromagnetic fields (EMF) associated with the use of mobile phones, particularly among young people, are increasing all the time. The potential deleterious effects of EMF exposure resulting from mobile phones being used in close proximity to the brain require particular evaluation. However, only a limited number of studies have investigated the effects of prenatal exposure to EMF in the development of the pyramidal cells using melatonin (MEL) and omega-3 (ω-3). We established seven groups of pregnant rats consisting of three animals each; control (CONT), SHAM, EMF, EMF + MEL, MEL, EMF + ω-3 and ω-3 alone. The rats in the EMF, EMF + MEL, EMF + ω-3 groups were exposed to 900 MHz EMF for 60 min/day in an exposure tube during the gestation period. The CONT, MEL and ω-3 group rats were not placed inside the exposure tube or exposed to EMF during the study period. After delivery, only spontaneously delivered male rat pups were selected for the establishment of further groups. Each group of offspring consisted of six animals. The optical fractionator technique was used to determine total pyramidal neuron numbers in the rat hippocampal region. The total number of pyramidal cells in the cornu ammonis (CA) in the EMF group was significantly lower than in the CONT, SHAM, EMF + MEL, and EMF + ω-3 groups. No significant difference was observed between the EMF, MEL and ω-3 groups. No difference was also observed between any groups in terms of rats' body or brain weights. MEL and ω-3 can protect the cell against neuronal damage in the hippocampus induced by 900 MHz EMF. However, further studies are now needed to evaluate the chronic effects of 900 MHz EMF on the brain in the prenatal period.

  4. Amino Acid Mixture Acts as a Potent VEGF Lowering Agent in CHO-K1 Cells Exposed to High Glucose.

    Science.gov (United States)

    Selvi, Radhakrishnan; Bhuvanasundar, Renganathan; Angayarkanni, Narayanasamy

    2017-04-01

    Though the role of amino acids in Diabetes Mellitus is controversial, the beneficial effect of amino acids in Diabetes Mellitus has been reported based on its anti-glycating property and insulin potentiating effects. In the current study, we evaluated the ROS generation and VEGF expression in CHO-K1 cells induced by high glucose concentration. The effect of amino acids treatment was studied under this condition to evaluate the VEGF lowering effect. CHO-K1 cells were treated various concentration of glucose (7 mmol, 17 mmol and 27 mmol) with and without free amino acids (5 mmol) or the amino acids mixture (AAM). Intracellular reactive oxygen species (ROS) was estimated by fluorescein dye (DCFDA), nitric oxide (NO) by Griess reaction, hydrogen peroxide (H2O2) by fluorimetry using Amplex red dye, super oxide dismutase (SOD) by spectrophotometry and VEGF by immunoblotting. High glucose condition significantly induced the expression of VEGF and this was reduced significantly by AAM treatment (p = 0.004). AAM also significantly decreased the cellular levels of ROS, NO, H2O2 as well as the SOD activity in CHO-K1 cells exposed to high glucose condition (p <0.05). The present study identified AAM as a potential VEGF lowering agent that intervenes at the level of oxidative stress in high glucose conditions as evaluated in CHO-K1 cells. Copyright © 2017 IMSS. Published by Elsevier Inc. All rights reserved.

  5. Matrix-degrading and pro-inflammatory changes in human vascular endothelial cells exposed to cigarette smoke condensate.

    Science.gov (United States)

    Nordskog, Brian K; Blixt, Allison D; Morgan, Walter T; Fields, Wanda R; Hellmann, Gary M

    2003-01-01

    Cigarette smoking has been associated with an increase in the severity and prevalence of atherosclerosis in the abdominal aorta. To begin our investigation of this finding, we used an integrated approach combining gene expression profiling, protein analysis, cytokine measurements, and cytotoxicity determinations to examine molecular responses of cultured human aortic and coronary endothelial cells exposed to cigarette smoke condensate (CSC) and nicotine. Exposure of endothelial cells to CSC (30 and 60 microg/mL TPM) for 24 h resulted in minimal cytotoxicity, and the upregulation of genes involved in matrix degradation (MMP-1, MMP-8, and MMP-9), xenobiotic metabolism (HO-1 and CYP1A2), and downregulation of genes involved in cell cycle regulation (including TOP2A, CCNB1, CCNA, CDKN3). Exposure of cells to a high physiological concentration of nicotine resulted in few differentially expressed genes. Immunoblot analysis of proteins selected from genes shown to be differentially regulated by microarray analysis revealed similar responses. Finally, a number of inflammatory cytokines measured in culture media were elevated in response to CSC. Together, these results describe a complex proinflammatory response, possibly mediating the recruitment of leukocytes through cytokine signaling. Additionally, fibrous cap destabilization may be facilitated by matrix metalloproteinase upregulation.

  6. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Chow, Paik Wah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Abdul Hamid, Zariyantey, E-mail: zyantey@ukm.edu.my [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Chan, Kok Meng [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Inayat-Hussain, Salmaan Hussain [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Rajab, Nor Fadilah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia)

    2015-04-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e{sup +} cells but reduced the total counts of Sca-1{sup +}, CD11b{sup +}, Gr-1{sup +}, and CD45{sup +} cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and

  7. Transcriptomics and methylomics of CD4-positive T cells in arsenic-exposed women.

    Science.gov (United States)

    Engström, Karin; Wojdacz, Tomasz K; Marabita, Francesco; Ewels, Philip; Käller, Max; Vezzi, Francesco; Prezza, Nicola; Gruselius, Joel; Vahter, Marie; Broberg, Karin

    2017-05-01

    Arsenic, a carcinogen with immunotoxic effects, is a common contaminant of drinking water and certain food worldwide. We hypothesized that chronic arsenic exposure alters gene expression, potentially by altering DNA methylation of genes encoding central components of the immune system. We therefore analyzed the transcriptomes (by RNA sequencing) and methylomes (by target-enrichment next-generation sequencing) of primary CD4-positive T cells from matched groups of four women each in the Argentinean Andes, with fivefold differences in urinary arsenic concentrations (median concentrations of urinary arsenic in the lower- and high-arsenic groups: 65 and 276 μg/l, respectively). Arsenic exposure was associated with genome-wide alterations of gene expression; principal component analysis indicated that the exposure explained 53% of the variance in gene expression among the top variable genes and 19% of 28,351 genes were differentially expressed (false discovery rate arsenic group. Arsenic exposure was associated with genome-wide DNA methylation; the high-arsenic group had 3% points higher genome-wide full methylation (>80% methylation) than the lower-arsenic group. Differentially methylated regions that were hyper-methylated in the high-arsenic group showed enrichment for immune-related gene ontologies that constitute the basic functions of CD4-positive T cells, such as isotype switching and lymphocyte activation and differentiation. In conclusion, chronic arsenic exposure from drinking water was related to changes in the transcriptome and methylome of CD4-positive T cells, both genome wide and in specific genes, supporting the hypothesis that arsenic causes immunotoxicity by interfering with gene expression and regulation.

  8. Reactive oxygen species regulated mitochondria-mediated apoptosis in PC12 cells exposed to chlorpyrifos

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    Lee, Jeong Eun [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Park, Jae Hyeon [Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of); Shin, In Chul [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Koh, Hyun Chul, E-mail: hckoh@hanyang.ac.kr [Department of Pharmacology, College of Medicine, Hanyang University, Seoul (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of)

    2012-09-01

    Reactive oxidative species (ROS) generated by environmental toxicants including pesticides could be one of the factors underlying the neuronal cell damage in neurodegenerative diseases. In this study we found that chlorpyrifos (CPF) induced apoptosis in dopaminergic neuronal components of PC12 cells as demonstrated by the activation of caspases and nuclear condensation. Furthermore, CPF also reduced the tyrosine hydroxylase-positive immunoreactivity in substantia nigra of the rat. In addition, CPF induced inhibition of mitochondrial complex I activity. Importantly, N-acetyl cysteine (NAC) treatment effectively blocked apoptosis via the caspase-9 and caspase-3 pathways while NAC attenuated the inhibition of mitochondrial complex I activity as well as the oxidative metabolism of dopamine (DA). These results demonstrated that CPF-induced apoptosis was involved in mitochondrial dysfunction through the production of ROS. In the response of cellular antioxidant systems to CPF, we found that CPF treatment increased HO-1 expression while the expression of CuZnSOD and MnSOD was reduced. In addition, we found that CPF treatment activated MAPK pathways, including ERK 1/2, the JNK, and the p38 MAP kinase in a time-dependent manner. NAC treatment abolished MAPK phosphorylation caused by CPF, indicating that ROS are upstream signals of MAPK. Interestingly, MAPK inhibitors abolished cytotoxicity and reduced ROS generation by CPF treatment. Our results demonstrate that CPF induced neuronal cell death in part through MAPK activation via ROS generation, suggesting its potential to generate oxidative stress via mitochondrial damage and its involvement in oxidative stress-related neurodegenerative disease. -- Highlights: ► Chlorpyrifos induces apoptosis. ► Chlorpyrifos inhibits mitochondrial complex I activity. ► ROS is involved in chlorpyrifos-induced apoptosis. ► Chlorpyrifos affects cellular antioxidant systems. ► Chlorpyrifos-induced apoptosis mediates activation of MAPK.

  9. Stress responses and conditioning effects in mesothelial cells exposed to peritoneal dialysis fluid.

    Science.gov (United States)

    Kratochwill, Klaus; Lechner, Michael; Siehs, Christian; Lederhuber, Hans C; Rehulka, Pavel; Endemann, Michaela; Kasper, David C; Herkner, Kurt R; Mayer, Bernd; Rizzi, Andreas; Aufricht, Christoph

    2009-04-01

    Renal replacement therapy by peritoneal dialysis is frequently complicated by technical failure. Peritoneal dialysis fluids (PDF) cause injury to the peritoneal mesothelial cell layer due to their cytotoxicity. As only isolated elements of the involved cellular processes have been studied before, we aimed at a global assessment of the mesothelial stress response to PDF. Following single or repeated exposure to PDF or control medium, proteomics and bioinformatics techniques were combined to study effects in mesothelial cells (MeT-5A). Protein expression was assessed by two-dimensional gel electrophoresis, and significantly altered spots were identified by MALDI-TOF MS and MS2 techniques. The lists of experimentally derived candidate proteins were expanded by a next neighbor approach and analyzed for significantly enriched biological processes. To address the problem of an unknown portion of false positive spots in 2DGE, only proteins showing significant p-values on both levels were further interpreted. Single PDF exposure resulted in reduction of biological processes in favor of reparative responses, including protein metabolism, modification and folding, with chaperones as a major subgroup. The observed biological processes triggered by this acute PDF exposure mainly contained functionally interwoven multitasking proteins contributing as well to cytoskeletal reorganization and defense mechanisms. Repeated PDF exposure resulted in attenuated protein regulation, reflecting inhibition of stress responses by high levels of preinduced chaperones. The identified proteins were less attributable to acute cellular injury but rather to specialized functions with a reduced number of involved multitasking proteins. This finding agrees well with the concept of conditioning effects and cytoprotection. In conclusion, this study describes the reprogrammed proteome of mesothelial cells during recovery from PDF exposure and adaption to repetitive stress. A broad stress response with

  10. Transcriptomic changes in mouse embryonic stem cells exposed to thalidomide during spontaneous differentiation

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    Xiugong Gao

    2015-09-01

    Full Text Available Thalidomide is a potent developmental toxicant that induces a range of birth defects, notably severe limb malformations. To unravel the molecular mechanisms underpinning the teratogenic effects of thalidomide, we used microarrays to study transcriptomic changes induced by thalidomide in an in vitro model based on the differentiation of mouse embryonic stem cells (mESCs, and published the major findings in a research article entitled “Thalidomide induced early gene expression perturbations indicative of human embryopathy in mouse embryonic stem cells” [1]. The data presented herein contains complementary information related to the aforementioned research article.

  11. Osteoblastic differentiation and stress response of human mesenchymal stem cells exposed to alternating current electric fields

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    Kaplan David L

    2011-01-01

    Full Text Available Abstract Background Electric fields are integral to many biological events, from maintaining cellular homeostasis to embryonic development to healing. The application of electric fields offers substantial therapeutic potential, while optimal dosing regimens and the underlying mechanisms responsible for the positive clinical impact are poorly understood. Methods The purpose of this study was to track the differentiation profile and stress response of human bone marrow derived mesenchymal stem cells (hMSCs undergoing osteogenic differentiation during exposure to a 20 mV/cm, 60 kHz electric field. Morphological and biochemical changes were imaged using endogenous two-photon excited fluorescence (TPEF and quantitatively assessed through eccentricity calculations and extraction of the redox ratio from NADH, FAD and lipofuscin contributions. Real time reverse transcriptase-polymerase chain reactions (RT-PCR were used to track osteogenic differentiation markers, namely alkaline phosphatase (ALP and collagen type 1 (col1, and stress response markers, such as heat shock protein 27 (hsp27 and heat shock protein 70 (hsp70. Comparisons of collagen deposition between the stimulated hMSCs and controls were examined through second harmonic generation (SHG imaging. Results Quantitative differences in cell morphology, as described through an eccentricity ratio, were found on days 2 and days 5 (p Conclusions Electrical stimulation is a useful tool to improve hMSC osteogenic differentiation, while heat shock proteins may reveal underlying mechanisms, and optical non-invasive imaging may be used to monitor the induced morphological and biochemical changes.

  12. [Cytogenetic investigations of bone marrow cells from mice exposed onboard biosatellite "Bion-M1"].

    Science.gov (United States)

    Dorozhkina, O V; Ivanov, A A

    2015-01-01

    The results of studying the mitotic activities and chromosomal aberrations in bone marrow cells from C57/BL6N mice with the help of the anaphase technique in 12 hours after completion of the 30-day "Bion-M1" mission and ground-based experiment using flight equipment are presented. A statistically reliable decline of the mitotic activity (0.74%) was found in cells taken from the space flown animals. In the ground-based experiment, a statistically reliable downward trend in proliferative activity (1.37%) was revealed after the comparison with groups of vivarium control (1.46-1.53%). In both experiments mice increased the number of initial mitotic phases (prophase + metaphase) relative to the sum of anaphases and telophases. The number of aberrant mitoses grew reliably in the group of flight animals by 29.7%, whereas in the ground-based experiment an upward trend was insignificant as their number increased up to 2.3% only. In the vivarium controls aberrant mitoses constituted 1.75-1.8%. An increase in chromosomal aberrations was largely due to such abnormalities as fragments. These findings seem to have been a result of summation of the effects of radiation and other stressful factors in space flight.

  13. Cigarette Smoke-Exposed Candida albicans Increased Chitin Production and Modulated Human Fibroblast Cell Responses

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    Humidah Alanazi

    2014-01-01

    Full Text Available The predisposition of cigarette smokers for development of respiratory and oral bacterial infections is well documented. Cigarette smoke can also contribute to yeast infection. The aim of this study was to investigate the effect of cigarette smoke condensate (CSC on C. albicans transition, chitin content, and response to environmental stress and to examine the interaction between CSC-pretreated C. albicans and normal human gingival fibroblasts. Following exposure to CSC, C. albicans transition from blastospore to hyphal form increased. CSC-pretreated yeast cells became significantly (P<0.01 sensitive to oxidation but significantly (P<0.01 resistant to both osmotic and heat stress. CSC-pretreated C. albicans expressed high levels of chitin, with 2- to 8-fold recorded under hyphal conditions. CSC-pretreated C. albicans adhered better to the gingival fibroblasts, proliferated almost three times more and adapted into hyphae, while the gingival fibroblasts recorded a significantly (P<0.01 slow growth rate but a significantly higher level of IL-1β when in contact with CSC-pretreated C. albicans. CSC was thus able to modulate both C. albicans transition through the cell wall chitin content and the interaction between C. albicans and normal human gingival fibroblasts. These findings may be relevant to fungal infections in the oral cavity in smokers.

  14. Epidermal Growth Factor-Like Growth Factors Prevent Apoptosis of Alcohol-Exposed Human Placental Cytotrophoblast Cells1

    Science.gov (United States)

    Wolff, Garen S.; Chiang, Po Jen; Smith, Susan M.; Romero, Roberto; Armant, D. Randall

    2007-01-01

    Maternal alcohol abuse during pregnancy can produce an array of birth defects comprising fetal alcohol syndrome. A hallmark of fetal alcohol syndrome is intrauterine growth retardation, which is associated with elevated apoptosis of placental cytotrophoblast cells. Using a human first trimester cytotrophoblast cell line, we examined the relationship between exposure to ethanol and cytotrophoblast survival, as well as the ameliorating effects of epidermal growth factor (EGF)-like growth factors produced by human cytotrophoblast cells. After exposure to 0–100 mM ethanol, cell death was quantified by the TUNEL method, and expression of the nuclear proliferation marker, Ki67, was measured by immunohistochemistry. The mode of cell death was determined by assessing annexin V binding, caspase 3 activation, pyknotic nuclear morphology, reduction of TUNEL by caspase inhibition, and cellular release of lactate dehydrogenase. Ethanol significantly reduced proliferation and increased cell death approximately 2.5-fold through the apoptotic pathway within 1–2 h of exposure to 50 mM alcohol. Exposure to 25–50 mM ethanol significantly increased transforming growth factor alpha (TGFA) and heparin-binding EGF-like growth factor (HBEGF), but not EGF or amphiregulin (AREG). When cytotrophoblasts were exposed concurrently to 100 mM ethanol and 1 nM HBEGF or TGFA, the increase in apoptosis was prevented, while EGF ameliorated at 10 nM and AREG was weakly effective. HBEGF survival-promoting activity required ligation of either of its cognate receptors, HER1 or HER4. These findings reveal the potential for ethanol to rapidly induce cytotrophoblast apoptosis. However, survival factor induction could provide cytotrophoblasts with an endogenous cytoprotective mechanism. PMID:17392498

  15. Caffeic Acid Reduces the Viability and Migration Rate of Oral Carcinoma Cells (SCC-25 Exposed to Low Concentrations of Ethanol

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    Arkadiusz Dziedzic

    2014-10-01

    Full Text Available Alcohol increases the risk of carcinoma originated from oral epithelium, but the biological effects of ultra-low doses of ethanol on existing carcinoma cells in combination with natural substances are still unclear. A role for ethanol (EtOH, taken in small amounts as an ingredient of some beverages or mouthwashes to change the growth behavior of established squamous cell carcinoma, has still not been examined sufficiently. We designed an in vitro study to determine the effect of caffeic acid (CFA on viability and migration ability of malignant oral epithelial keratinocytes, exposed to ultra-low concentrations (maximum 100 mmol/L EtOH. MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-dimethyltetrazolium bromide and LDH (lactate dehydrogenase assays were used to assess the cytotoxic effect of EtOH/CFA and the viability of squamous carcinoma SCC-25 cells (ATCC CRL-1628, mobile part of the tongue. Tested EtOH concentrations were: 2.5, 5, 10, 25, 50, and 100 mmol/L, along with an equal CFA concentration of 50 μmol/L. Carcinoma cells’ migration was investigated by monolayer “wound” healing assay. We demonstrated that very low concentrations of EtOH ranging between 2.5 and 10 mmol/L may induce the viability of oral squamous cell carcinoma cells, while the results following addition of CFA reveal an antagonistic effect, attenuating pro-proliferative EtOH activity. The migration rate of oral squamous carcinoma cells can be significantly inhibited by the biological activity of caffeic acid.

  16. Investigating longitudinal changes in the mechanical properties of MCF-7 cells exposed to paclitaxol using particle tracking microrheology

    Science.gov (United States)

    El Kaffas, Ahmed; Bekah, Devesh; Rui, Min; Kumaradas, J. Carl; Kolios, Michael C.

    2013-02-01

    Evidence suggests that compression and shear wave elastography are sensitive to the mechanical property changes occuring in dying cells following chemotherapy, and can hence be used to monitor cancer treatment response. A qualitative and quantitative understanding of the mechanical changes at the cellular level would allow to better infer how these changes affect macroscopic tissue mechanical properties and therefore allow the optimization of elastographic techniques (such as shear wave elastography) for the monitoring of cancer therapy. We used intracellular particle tracking microrheology (PTM) to investigate the mechanical property changes of cells exposed to paclitaxol, a mitotic inhibitor used in cancer chemotherapy. The average elastic and viscous moduli of the cytoplasm of treated MCF-7 breast cancer cells were calculated for frequency ranges between 0.2 and 100 rad s-1 (corresponding to 0.03 and 15.92 Hz, respectively). A significant increase in the complex shear modulus of the cell cytoplasm was detected at 12 h post treatment. At 24 h after drug exposure, the elastic and viscous moduli increased by a total of 191.3 Pa (>8000×) and 9 Pa (˜9×), respectively for low frequency shear modulus measurements (at 1 rad s-1). At higher frequencies (10 rad s-1), the elastic and viscous moduli increased by 188.5 Pa (˜60×) and 1.7 Pa (˜1.1×), respectively. Our work demonstrates that PTM can be used to measure changes in the mechanical properties of treated cells and that cell elasticity significantly increases by 24 h after chemotherapy exposure.

  17. Decline of FoxP3+ Regulatory CD4 T Cells in Peripheral Blood of Children Heavily Exposed to Malaria.

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    Michelle J Boyle

    2015-07-01

    Full Text Available FoxP3+ regulatory CD4 T cells (Tregs help to maintain the delicate balance between pathogen-specific immunity and immune-mediated pathology. Prior studies suggest that Tregs are induced by P. falciparum both in vivo and in vitro; however, the factors influencing Treg homeostasis during acute and chronic infections, and their role in malaria immunopathogenesis, remain unclear. We assessed the frequency and phenotype of Tregs in well-characterized cohorts of children residing in a region of high malaria endemicity in Uganda. We found that both the frequency and absolute numbers of FoxP3+ Tregs in peripheral blood declined markedly with increasing prior malaria incidence. Longitudinal measurements confirmed that this decline occurred only among highly malaria-exposed children. The decline of Tregs from peripheral blood was accompanied by reduced in vitro induction of Tregs by parasite antigen and decreased expression of TNFR2 on Tregs among children who had intense prior exposure to malaria. While Treg frequencies were not associated with protection from malaria, there was a trend toward reduced risk of symptomatic malaria once infected with P. falciparum among children with lower Treg frequencies. These data demonstrate that chronic malaria exposure results in altered Treg homeostasis, which may impact the development of antimalarial immunity in naturally exposed populations.

  18. Effect of exposure time, particle size and uptake pathways in immune cell lysosomal cytotoxicity of mussels exposed to silver nanoparticles.

    Science.gov (United States)

    Bouallegui, Younes; Ben Younes, Ridha; Turki, Faten; Mezni, Amine; Oueslati, Ridha

    2018-04-01

    Cytotoxicity evaluation of hemocytes (lysosomal membrane stability [LMS] assay) from Mytilus galloprovincialis Lamarck, exposed to a sublethal dose (100 μg/L) of two size of silver nanoparticles (AgNPs: <50 nm and <100 nm) - prior to and after inhibition of potential uptake pathways (i.e., clathrin- and caveolae-mediated endocytosis) within different times of exposure (3, 6, 12 h) - showed that there was a significant cytotoxic effect on immune cells of mussels exposed for different times to either AgNP size (p < 0.01); the greater effect was with the smaller size. However, hemocytes seemed more sensitive to the larger AgNP after clathrin-mediated endocytosis was blocked (p < 0.01); this was not so with inhibition of caveolae-mediated endocytosis. Dimethyl-sulfoxide (DMSO) did not impart a carrier-mediated effect despite an enhanced cytotoxicity when DMSO was present with AgNP. From these results, it is concluded that the immunotoxicity of AgNP in mussels was size-dependent as well as length of exposure-dependent. It was also clear that nanoparticles (NP) internalization mechanisms were a major factor underlying any toxicity.

  19. Nuclear survivin and its relationship to DNA damage repair genes in non-small cell lung cancer investigated using tissue array.

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    Songliu Hu

    Full Text Available To investigate the predictive role and association of nuclear survivin and the DNA double-strand breaks repair genes in non-small cell lung cancer (NSCLC: DNA-dependent protein kinase catalytic subunit (DNA-PKcs, Ku heterodimeric regulatory complex 70-KD subunit (Ku70 and ataxia-telangiectasia mutated (ATM.The protein expression of nuclear survivin, DNA-PKcs, Ku70 and ATM were investigated using immunohistochemistry in tumors from 256 patients with surgically resected NSCLC. Furthermore, we analyzed the correlation between the expression of nuclear survivin, DNA-PKcs, Ku70 and ATM. Univariate and multivariate analyses were performed to determine the prognostic factors that inuenced the overall survival and disease-free survival of NSCLC.The expression of nuclear survivin, DNA-PKcs, Ku70 and ATM was significantly higher in tumor tissues than in normal tissues. By dichotomizing the specimens as expressing low or high levels of nuclear survivin, nuclear survivin correlated significantly with the pathologic stage (P = 0.009 and lymph node status (P = 0.004. The nuclear survivin levels were an independent prognostic factor for both the overall survival and the disease-free survival in univariate and multivariate analyses. Patients with low Ku70 and DNA-PKcs expression had a greater benefit from radiotherapy than patients with high expression of Ku70 (P = 0.012 and DNA-PKcs (P = 0.02. Nuclear survivin expression positively correlated with DNA-PKcs (P<0.001 and Ku70 expression (P<0.001.Nuclear survivin may be a prognostic factor for overall survival in patients with resected stage I-IIIA NSCLC. DNA-PKcs and Ku70 could predict the effect of radiotherapy in patients with NSCLC. Nuclear survivin may also stimulates DNA double-strand breaks repair by its interaction with DNA-PKcs and Ku70.

  20. p38 MAPK-Mediated Bmi-1 Down-Regulation and Defective Proliferation in ATM-Deficient Neural Stem Cells Can Be Restored by Akt Activation

    Science.gov (United States)

    Kim, Jeesun; Hwangbo, Jeon; Wong, Paul K. Y.

    2011-01-01

    A-T (ataxia telangiectasia) is a genetic disease caused by a mutation in the Atm (A-T mutated) gene that leads to neurodegeneration. Despite an increase in the numbers of studies in this area in recent years, the mechanisms underlying neurodegeneration in human A-T are still poorly understood. Previous studies demonstrated that neural stem cells (NSCs) isolated from the subventricular zone (SVZ) of Atm-/- mouse brains show defective self-renewal and proliferation, which is accompanied by activation of chronic p38 mitogen-activated protein kinase (MAPK) and a lower level of the polycomb protein Bmi-1. However, the mechanism underlying Bmi-1 down-regulation and its relevance to defective proliferation in Atm-/- NSCs remained unclear. Here, we show that over-expression of Bmi-1 increases self-renewal and proliferation of Atm-/- NSCs to normal, indicating that defective proliferation in Atm-/- NSCs is a consequence of down-regulation of Bmi-1. We also demonstrate that epidermal growth factor (EGF)-induced Akt phosphorylation renders Bmi-1 resistant to the proteasomal degradation, leading to its stabilization and accumulation in the nucleus. However, inhibition of the Akt-dependent Bmi-1 stabilizing process by p38 MAPK signaling reduces the levels of Bmi-1. Treatment of the Atm-/- NSCs with a specific p38 MAPK inhibitor SB203580 extended Bmi-1 posttranscriptional turnover and H2A ubiquitination in Atm-/- NSCs. Our observations demonstrate the molecular basis underlying the impairment of self-renewal and proliferation in Atm-/- NSCs through the p38 MAPK-Akt-Bmi-1-p21 signaling pathway. PMID:21305053

  1. The telomeric protein TRF2 is critical for the protection of A549 cells from both telomere erosion and DNA double-strand breaks driven by salvicine.

    Science.gov (United States)

    Zhang, Yong-Wei; Zhang, Zhi-Xiang; Miao, Ze-Hong; Ding, Jian

    2008-03-01

    Telomere repeat binding factor 2 (TRF2) has been increasingly recognized to be involved in DNA damage response and telomere maintenance. Our previous report found that salvicine (SAL), a novel topoisomerase II poison, elicited DNA double-strand breaks and telomere erosion in separate experimental systems. However, it remains to be clarified whether they share a common response to these two events and in particular whether TRF2 is involved in this process. In this study, we found that SAL concurrently induced DNA double-strand breaks, telomeric DNA damage, and telomere erosion in lung carcinoma A549 cells. It was unexpected to find that SAL led to disruption of TRF2, independently of either its transcription or proteasome-mediated degradation. By overexpressing the full-length trf2 gene and transfecting TRF2 small interfering RNAs, we showed that TRF2 protein protected both telomeric and genomic DNA from the SAL-elicited events. It is noteworthy that although both the Ataxia-telangiectasia-mutated (ATM) and the ATM- and Rad3-related (ATR) kinases responded to the SAL-induced DNA damages, only ATR was essential for the telomere erosion. The study also showed that the activated ATR augmented the SAL-triggered TRF2 disruption, whereas TRF2 reduction in turn enhanced ATR function. All of these findings suggest the emerging significance of TRF2 protecting both telomeric DNA and genomic DNA on the one hand and reveal the mutual modulation between ATR and TRF2 in sensing DNA damage signaling during cancer development on the other hand.

  2. Genomic Profiling of a Human Leukemic Monocytic Cell-Line (THP-1 Exposed to Alpha Particle Radiation

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    Vinita Chauhan

    2012-01-01

    Full Text Available This study examined alpha (α- particle radiation effects on global changes in gene expression in human leukemic monocytic cells (THP-1 for the purposes of mining for candidate biomarkers that could be used for the development of a biological assessment tool. THP-1 cells were exposed to α-particle radiation at a dose range of 0 to 1.5 Gy. Twenty-four hours and three days after exposure gene expression was monitored using microarray technology. A total of 16 genes were dose responsive and classified as early onset due to their expression 24 h after exposure. Forty-eight transcripts were dose responsive and classified as late-onset as they were expressed 72 h after exposure. Among these genes, 6 genes were time and dose responsive and validated further using alternate technology. These transcripts were upregulated and associated with biological processes related to immune function, organelle stability and cell signalling/communication. This panel of genes merits further validation to determine if they are strong candidate biomarkers indicative of α-particle exposure.

  3. The effect of citrus flavanones on the redox homeostasis in cells exposed to oxidative stress – studies in vitro

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    Ewa Kurzeja

    2016-06-01

    Full Text Available ioxidants in citrus fruits are beneficial for health, which is connected with their anti-inflammatory, anti-atherogenic and anti-carcinogenic properties. The present study was undertaken to investigate whether – and in what way – the presence of flavanones influences the redox homeostasis of fibroblasts and alleviates the effects of oxidative stress. Material and methods: The study was conducted on murine fibroblast cell cultures with the addition of flavanones (hesperidin, hesperetin, naringin, naringenin, exposed to oxidative stress (Fe/Asc. In cell homogenates, the activity of superoxide dismutase (SOD and glutathione peroxidase (GPx was measured; in the medium, the concentration of nitric oxide was measured. Results and conclusion: Our results demonstrate that the addition of naringenin, hesperetin, naringin and hesperidin has a protective effect on cells subjected to oxidative stress The changes observed are particularly visible in the case of aglycone forms of both compounds. Despite the protective properties against oxidative stress which flavanones display, we determined distrubances in redox homeostasis in comparison to the control culture.

  4. ER-Dependent Ca++-mediated Cytosolic ROS as an Effector for Induction of Mitochondrial Apoptotic and ATM-JNK Signal Pathways in Gallic Acid-treated Human Oral Cancer Cells.

    Science.gov (United States)

    Lu, Yao-Cheng; Lin, Meng-Liang; Su, Hong-Lin; Chen, Shih-Shun

    2016-02-01

    Release of calcium (Ca(++)) from the endoplasmic reticulum (ER) has been proposed to be involved in induction of apoptosis by oxidative stress. Using inhibitor of ER Ca(++) release dantrolene and inhibitor of mitochondrial Ca(++) uptake Ru-360, we demonstrated that Ca(++) release from the ER was associated with generation of reactive oxygen species (ROS), loss of mitochondrial membrane potential, and apoptosis of human oral cancer (OC) cells induced by gallic acid (GA). Small interfering RNA-mediated suppression of protein kinase RNA-like endoplasmic reticulum kinase inhibited tunicamycin-induced induction of 78 kDa glucose-regulated protein, C/EBP homologous protein, pro-caspase-12 cleavage, cytosolic Ca(++) increase and apoptosis, but did not attenuate the increase in cytosolic Ca(++) level and apoptosis induced by GA. Ataxia telangiectasia mutated (ATM)-mediated c-Jun N-terminal kinase (JNK) phosphorylation and apoptosis by GA was blocked by dantrolene. The specificity of ROS-mediated ATM-JNK activation was confirmed by treatment with N-acetylcysteine, a ROS scavenger. Blockade of ATM activation by specific inhibitor KU55933, short hairpin RNA, or kinase-dead ATM overexpression suppressed JNK phosphorylation but did not completely inhibit cytosolic ROS production, mitochondrial cytochrome c release, pro-caspase-3 cleavage, and apoptosis induced by GA. Taken together, these results indicate that GA induces OC cell apoptosis by inducing the activation of mitochondrial apoptotic and ATM-JNK signal pathways, likely through ER Ca(++)-mediated ROS production. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  5. Memory B cells are a more reliable archive for historical antimalarial responses than plasma antibodies in no-longer exposed children.

    Science.gov (United States)

    Ndungu, Francis Maina; Olotu, Ally; Mwacharo, Jedidah; Nyonda, Mary; Apfeld, Jordan; Mramba, Lazarus K; Fegan, Gregory W; Bejon, Philip; Marsh, Kevin

    2012-05-22

    Humans respond to foreign antigen by generating plasma Abs and memory B cells (MBCs). The Ab response then declines, sometimes to below the limit of detection. In contrast, MBCs are generally thought to be long-lived. We tested and compared Plasmodium falciparum (Pf)-specific Ab and MBC responses in two populations of children: (i) previously exposed children who had documented Pf infections several years ago, but minimal exposure since then; and (ii) persistently exposed children living in a separate but nearby endemic area. We found that although Pf-specific plasma Abs were lower in previously exposed children compared with persistently exposed children, their cognate MBCs were maintained at similar frequencies. We conclude that serological analysis by itself would greatly underestimate the true memory of Pf-specific Ab responses in previously exposed children living in areas where Pf transmission has been reduced or eliminated.

  6. Sodium nitrite induces acute central nervous system toxicity in guinea pigs exposed to systemic cell-free hemoglobin

    Energy Technology Data Exchange (ETDEWEB)

    Buehler, Paul W.; Butt, Omer I. [Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); D' Agnillo, Felice, E-mail: felice.dagnillo@fda.hhs.gov [Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States)

    2011-06-10

    Highlights: {yields} Toxicological implications associated with the use of NaNO{sub 2} therapy to treat systemic cell-free Hb exposure are not well-defined. {yields} Systemic Hb exposure followed by NaNO{sub 2} infusion induces acute CNS toxicities in guinea pigs. {yields} These CNS effects were not reproduced by the infusion of cell-free Hb or NaNO{sub 2} alone. {yields} NaNO{sub 2}-mediated oxidation of cell-free Hb may play a causative role in the observed CNS changes. -- Abstract: Systemic cell-free hemoglobin (Hb) released via hemolysis disrupts vascular homeostasis, in part, through the scavenging of nitric oxide (NO). Sodium nitrite (NaNO{sub 2}) therapy can attenuate the hypertensive effects of Hb. However, the chemical reactivity of NaNO{sub 2} with Hb may enhance heme- or iron-mediated toxicities. Here, we investigate the effect of NaNO{sub 2} on the central nervous system (CNS) in guinea pigs exposed to systemic cell-free Hb. Intravascular infusion of NaNO{sub 2}, at doses sufficient to alleviate Hb-mediated blood pressure changes, reduced the expression of occludin, but not zona occludens-1 (ZO-1) or claudin-5, in cerebral tight junctions 4 h after Hb infusion. This was accompanied by increased perivascular heme oxygenase-1 expression, neuronal iron deposition, increased astrocyte and microglial activation, and reduced expression of neuron-specific nuclear protein (NeuN). These CNS changes were not observed in animals treated with Hb or NaNO{sub 2} alone. Taken together, these findings suggest that the use of nitrite salts to treat systemic Hb exposure may promote acute CNS toxicity.

  7. Effects of Carbocysteine and Beclomethasone on Histone Acetylation/Deacetylation Processes in Cigarette Smoke Exposed Bronchial Epithelial Cells.

    Science.gov (United States)

    Pace, Elisabetta; Di Vincenzo, Serena; Ferraro, Maria; Siena, Liboria; Chiappara, Giuseppina; Dino, Paola; Vitulo, Patrizio; Bertani, Alessandro; Saibene, Federico; Lanata, Luigi; Gjomarkaj, Mark

    2017-10-01

    Histone deacetylase expression/activity may control inflammation, cell senescence, and responses to corticosteroids. Cigarette smoke exposure, increasing oxidative stress, may negatively affect deacetylase expression/activity. The effects of cigarette smoke extracts (CSE), carbocysteine, and beclomethasone dipropionate on chromatin remodeling processes in human bronchial epithelial cells are largely unknown. The present study was aimed to assess the effects of cigarette smoke, carbocysteine, and beclomethasone dipropionate on histone deacetylase 3 (HDAC3) expression/activity, N-CoR (nuclear receptor corepressor) expression, histone acetyltransferases (HAT) (p300/CBP) expression, p-CREB and IL-1 m-RNA expression, neutrophil chemotaxis. Increased p-CREB expression was observed in the bronchial epithelium of smokers. CSE increased p-CREB expression and decreased HDAC3 expression and activity and N-CoR m-RNA and protein expression. At the same time, CSE increased the expression of the HAT, p300/CBP. All these events increased acetylation processes within the cells and were associated to increased IL-1 m-RNA expression and neutrophil chemotaxis. The incubation of CSE exposed cells with carbocysteine and beclomethasone counteracted the effects of cigarette smoke on HDAC3 and N-CoR but not on p300/CBP. The increased deacetylation processes due to carbocysteine and beclomethasone dipropionate incubation is associated to reduced p-CREB, IL-1 m-RNA expression, neutrophil chemotaxis. These findings suggest a new role of combination therapy with carbocysteine and beclomethasone dipropionate in restoring deacetylation processes compromised by cigarette smoke exposure. J. Cell. Physiol. 232: 2851-2859, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  8. DNA hypermethylation of CD3(+) T cells from cord blood of infants exposed to intrauterine growth restriction.

    Science.gov (United States)

    Williams, Lyda; Seki, Yoshinori; Delahaye, Fabien; Cheng, Alex; Fuloria, Mamta; Hughes Einstein, Francine; Charron, Maureen J

    2016-08-01

    Intrauterine growth restriction (IUGR) is associated with increased susceptibility to obesity, metabolic syndrome and type 2 diabetes. Although the mechanisms underlying the developmental origins of metabolic disease are poorly understood, evidence suggests that epigenomic alterations play a critical role. We sought to identify changes in DNA methylation patterns that are associated with IUGR in CD3(+) T cells purified from umbilical cord blood obtained from male newborns who were appropriate for gestational age (AGA) or who had been exposed to IUGR. CD3(+) T cells were isolated from cord blood obtained from IUGR and AGA infants. The genome-wide methylation profile in eight AGA and seven IUGR samples was determined using the HELP tagging assay. Validation analysis using targeted bisulfite sequencing and bisulfite massARRAY was performed on the original cohort as well as biological replicates consisting of two AGA and four IUGR infants. The Segway algorithm was used to identify methylation changes within regulatory regions of the genome. A global shift towards hypermethylation in IUGR was seen compared with AGA (89.8% of 4,425 differentially methylated loci), targeted to regulatory regions of the genome, specifically promoters and enhancers. Pathway analysis identified dysregulation of pathways involved in metabolic disease (type 2 diabetes mellitus, insulin signalling, mitogen-activated protein kinase signalling) and T cell development, regulation and activation (T cell receptor signalling), as well as transcription factors (TCF3, LEF1 and NFATC) that regulate T cells. Furthermore, bump-hunting analysis revealed differentially methylated regions in PRDM16 and HLA-DPB1, genes important for adipose tissue differentiation, stem cell maintenance and function and T cell activation. Our findings suggest that the alterations in methylation patterns observed in IUGR CD3(+) T cells may have functional consequences in targeted genes, regulatory regions and transcription

  9. Live-Cell Imaging Visualizes Frequent Mitotic Skipping During Senescence-Like Growth Arrest in Mammary Carcinoma Cells Exposed to Ionizing Radiation

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Masatoshi, E-mail: msuzuki@nagasaki-u.ac.jp [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan); Yamauchi, Motohiro; Oka, Yasuyoshi; Suzuki, Keiji; Yamashita, Shunichi [Department of Radiation Medical Sciences, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki (Japan)

    2012-06-01

    Purpose: Senescence-like growth arrest in human solid carcinomas is now recognized as the major outcome of radiotherapy. This study was designed to analyze cell cycle during the process of senescence-like growth arrest in mammary carcinoma cells exposed to X-rays. Methods and Materials: Fluorescent ubiquitination-based cell cycle indicators were introduced into the human mammary carcinoma cell line MCF-7. Cell cycle was sequentially monitored by live-cell imaging for up to 5 days after exposure to 10 Gy of X-rays. Results: Live-cell imaging revealed that cell cycle transition from G2 to G1 phase without mitosis, so-called mitotic skipping, was observed in 17.1% and 69.8% of G1- and G2-irradiated cells, respectively. Entry to G1 phase was confirmed by the nuclear accumulation of mKO{sub 2}-hCdt1 as well as cyclin E, which was inversely correlated to the accumulation of G2-specific markers such as mAG-hGeminin and CENP-F. More than 90% of cells skipping mitosis were persistently arrested in G1 phase and showed positive staining for the senescent biochemical marker, which is senescence-associated ss-galactosidase, indicating induction of senescence-like growth arrest accompanied by mitotic skipping. While G2 irradiation with higher doses of X-rays induced mitotic skipping in approximately 80% of cells, transduction of short hairpin RNA (shRNA) for p53 significantly suppressed mitotic skipping, suggesting that ionizing radiation-induced mitotic skipping is associated with p53 function. Conclusions: The present study found the pathway of senescence-like growth arrest in G1 phase without mitotic entry following G2-irradiation.

  10. Investigation of toxic factors affecting cells of rat brains exposed to 3-methylcatechol

    Directory of Open Access Journals (Sweden)

    George Emílio Sampaio Barreto

    2007-09-01

    Full Text Available The aim of this work was to study the effects of 3MC on the peroxidation of biomolecules in nuclear fractions and nonsynaptic mitochondrial respiration in organelles obtained from rat brains. The cytotoxicity towards rat primary astrocytes in vitro was also tested. 3MC at 1mM oxidized consuming oxygen at a rate of 1.98 ± 0.19 µM.min-1 and formed reactive quinones. At the same concentration, 3MC induced peroxidation of biomolecules in nuclear fractions obtained from rat brain homogenates and inhibited state 2 FADH2-linked respiration in nonsynaptic mitochondria. Furthermore, 3MC oxidized in the culture medium, leading to the formation of quinones. This toluene metabolite was cytotoxic to rat primary astrocytes. The concentration that killed 50% of cells after 72 h was 107 mM. The results of the study indicated a direct relationship between cytotoxicity and 3MC oxidation.O 3-metilcatecol (3MC é um metabólito do tolueno. Para esclarecer se o 3MC seria tóxico para o sistema nervoso central, examinou-se seus efeitos sobre a peroxidação de biomoléculas em frações nucleares e a respiração mitocondrial em organelas obtidas de cérebros de ratos. Também se testou a citotoxicidade para astrócitos primários de ratos. O 3MC a 1mM oxida-se consumindo oxigênio a uma taxa de 1,98 ± 0,19 mM.min-1, formando quinonas reativas. Nessa mesma concentração o 3MC peroxidou biomoléculas nas frações nucleares. Esse composto também inibiu o estado 2 da respiração mitocondrial associada ao FADH2. Além disso, o 3MC também se oxida em meio de cultura levando à formação de quinonas. Esse metabólito do tolueno foi citotóxico para astrócitos de ratos. A concentração que matou 50% das células após 72 horas foi 107 mM. Os resultados desse estudo indicam uma relação direta entre a citotoxicidade e a oxidação do 3MC.

  11. Cytogenetic effects in bone marrow cells of mice exposed on the biosatellite "BION-M1"

    Science.gov (United States)

    Dorozhkina, Olga; Ivanov, Alexander

    In studies of cytogenetic damage in blood lymphocytes of astronauts, conducted in recent years, have shown an increase in the frequency of chromosomal damage bound, as believe, with influence on an organism of astronauts of space radiation (B.S. Fedorenko, G.P. Snigireva, 2004). However, in recent years published evidence that both acute and chronic stress induce chromosomal aberrations and modified genome sensitivity to mutagens of different nature, including to ionizing radiation (F.I. Ingel et al, 2005 ). This question is especially actual for space biology and medicine due to a number of specific features of space flights, when the interaction of factors more pronounced than in normal terrestrial conditions. In experiment "BION - M1" by anaphase method was determined level of chromosomal aberrations in bone marrow cells of tibia of mice. Flight duration biosatellite "BION - M1" was 30 days in Earth orbit. Euthanasia of experimental animals was carried out at intervals of 15-20 minutes by method of cervical dislocation after 12 hours from the moment of landing satellite. Level of chromosomal aberrations in vivarium-housed control mice was 1,75 ± 0,6% and 1,8 ± 0,45%, while the mitotic index 1,46 ± 0,09% and 1,53 ± 0,05%. Differences are not significant. The maintenance of animals in experiment with the onboard equipment (ground experiment) led to some increase in aberrant mitoses (2,3 ± 0,4%) and to decrease in a mitotic index (1,37 ± 0,02%). In the flight experiment "BION - M1" statistically significant increase of level of chromosomal aberrations (29,7 ± 4,18%) and a decrease in the mitotic index (0,74 ± 0,07%). Since the mouse is a suitable experimental model , also had several ground experiments on research of combined effect of irradiation and other stress factors specific to space flight, with marked tendency to increase the level of aberrant mitoses under the combined action of radiation and stress exposure group housing male mice. Statistically

  12. Antibacterial and antigelatinolytic effects of Satureja hortensis L. essential oil on epithelial cells exposed to Fusobacterium nucleatum.

    Science.gov (United States)

    Zeidán-Chuliá, Fares; Keskin, Mutlu; Könönen, Eija; Uitto, Veli-Jukka; Söderling, Eva; Moreira, José Cláudio Fonseca; Gürsoy, Ulvi K

    2015-04-01

    The present report examined the effects of essential oils (EOs) from Satureja hortensis L. and Salvia fruticosa M. on the viability and outer membrane permeability of the periodontopathogen Fusobacterium nucleatum, a key bacteria in oral biofilms, as well as the inhibition of matrix metalloproteinase (MMP-2 and MMP-9) activities in epithelial cells exposed to such bacteria. Membrane permeability was tested by measuring the N-phenyl-1-naphthylamine uptake and bacterial viability by using the commercially available Live/Dead BacLight kit. In addition, gelatin zymography was performed to analyze the inhibition of F. nucleatum-induced MMP-2 and MMP-9 activities in HaCaT cells. We showed that 5, 10, and 25 μL/mL of Sat. hortensis L. EO decreased the ratio of live/dead bacteria and increased the outer membrane permeability in a range of time from 0 to 5 min. Treatments with 10 and 25 μL/mL of Sal. fruticosa M. also increased the membrane permeability and 5, 10, and 25 μL/mL of both EOs inhibited MMP-2 and MMP-9 activities in keratinocytes induced after exposure of 24 h to F. nucleatum. We conclude that antibacterial and antigelatinolytic activities of Sat. hortensis L. EO have potential for the treatment of periodontal inflammation.

  13. Changes of inflammatory cells in rat lungs exposed to diesel emissions; Diesel haiki bakuro ni yoru rat hai no ensho saibo no henka

    Energy Technology Data Exchange (ETDEWEB)

    Kato, A. [Japan Automobile Research Institute Inc., Tsukuba (Japan); Kagawa, J. [Tokyo Women`s Medical College, Tokyo (Japan)

    1998-05-01

    Study was made on the effect of exposure to diesel emissions on inflammatory cells in a rat lungs. Four kinds of exposure gases with different contents of NO2 and particulate were prepared by diluting diesel emissions. Rats were exposed to diluted diesel emissions for 24 months, and inflammatory cells were detected morphologically in light microscopic and TEM specimens. As a result, particle-laden- alveolar macrophages increased dose- and time-dependently into the submucosa of intrapulmonary bronchioles, alveolar spaces and interstitume of alveolar walls, and bronchoassociated lymphatic tissues. Mast cells infiltrated into the interspaces of epithelial cells in airways. In the submucosa of the terminal bronchioles and the interstitume of alveolar walls, some sorts of inflammatory cells such as mast cells, plasma cells, neutrophils and lymphocytes infiltrated, and some cells showed cell-to-cell contacts. However, the airways were rarely injured by infiltration of inflammatory cells except for a fibrotic change. 2 refs., 2 figs., 2 tabs.

  14. Comprehensive analysis of 5-aminolevulinic acid dehydrogenase (ALAD variants and renal cell carcinoma risk among individuals exposed to lead.

    Directory of Open Access Journals (Sweden)

    Dana M van Bemmel

    Full Text Available BACKGROUND: Epidemiologic studies are reporting associations between lead exposure and human cancers. A polymorphism in the 5-aminolevulinic acid dehydratase (ALAD gene affects lead toxicokinetics and may modify the adverse effects of lead. METHODS: The objective of this study was to evaluate single-nucleotide polymorphisms (SNPs tagging the ALAD region among renal cancer cases and controls to determine whether genetic variation alters the relationship between lead and renal cancer. Occupational exposure to lead and risk of cancer was examined in a case-control study of renal cell carcinoma (RCC. Comprehensive analysis of variation across the ALAD gene was assessed using a tagging SNP approach among 987 cases and 1298 controls. Occupational lead exposure was estimated using questionnaire-based exposure assessment and expert review. Odds ratios (OR and 95% confidence intervals (CI were calculated using logistic regression. RESULTS: The adjusted risk associated with the ALAD variant rs8177796(CT/TT was increased (OR = 1.35, 95%CI = 1.05-1.73, p-value = 0.02 when compared to the major allele, regardless of lead exposure. Joint effects of lead and ALAD rs2761016 suggest an increased RCC risk for the homozygous wild-type and heterozygous alleles ((GGOR = 2.68, 95%CI = 1.17-6.12, p = 0.01; (GAOR = 1.79, 95%CI = 1.06-3.04 with an interaction approaching significance (p(int = 0.06. No significant modification in RCC risk was observed for the functional variant rs1800435(K68N. Haplotype analysis identified a region associated with risk supporting tagging SNP results. CONCLUSION: A common genetic variation in ALAD may alter the risk of RCC overall, and among individuals occupationally exposed to lead. Further work in larger exposed populations is warranted to determine if ALAD modifies RCC risk associated with lead exposure.

  15. Discovery of novel biomarkers by microarray analysis of peripheral blood mononuclear cell gene expression in benzene-exposed workers.

    Science.gov (United States)

    Forrest, Matthew S; Lan, Qing; Hubbard, Alan E; Zhang, Luoping; Vermeulen, Roel; Zhao, Xin; Li, Guilan; Wu, Yen-Ying; Shen, Min; Yin, Songnian; Chanock, Stephen J; Rothman, Nathaniel; Smith, Martyn T

    2005-06-01

    Benzene is an industrial chemical and component of gasoline that is an established cause of leukemia. To better understand the risk benzene poses, we examined the effect of benzene exposure on peripheral blood mononuclear cell (PBMC) gene expression in a population of shoe-factory workers with well-characterized occupational exposures using microarrays and real-time polymerase chain reaction (PCR). PBMC RNA was stabilized in the field and analyzed using a comprehensive human array, the U133A/B Affymetrix GeneChip set. A matched analysis of six exposed-control pairs was performed. A combination of robust multiarray analysis and ordering of genes using paired t-statistics, along with bootstrapping to control for a 5% familywise error rate, was used to identify differentially expressed genes in a global analysis. This resulted in a set of 29 known genes being identified that were highly likely to be differentially expressed. We also repeated these analyses on a smaller subset of 508 cytokine probe sets and found that the expression of 19 known cytokine genes was significantly different between the exposed and the control subjects. Six genes were selected for confirmation by real-time PCR, and of these, CXCL16, ZNF331, JUN, and PF4 were the most significantly affected by benzene exposure, a finding that was confirmed in a larger data set from 28 subjects. The altered expression was not caused by changes in the makeup of the PBMC fraction. Thus, microarray analysis along with real-time PCR confirmation reveals that altered expressions of CXCL16, ZNF331, JUN, and PF4 are potential biomarkers of benzene exposure.

  16. Allele and genotype frequency of a genetic variant in ataxia telangiectasia mutated gene affecting glycemic response to metformin in South Indian population

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    Saranya Vilvanathan

    2014-01-01

    Full Text Available Allele and genotype frequency of a genetic variant in ATM gene affecting glycemic response to metformin in South Indian population . Context: The novel polymorphism in ATM gene (rs11212617, which is implicated to have association with metformin response, exhibits inter-ethnic variability in the allele and genotype frequency distribution . Aims and Design: The objective of the present study is to establish the allele and genotype frequency of rs11212617 single nucleotide polymorphism in ATM gene, in South Indian population and to find if this variant has any role in the etiology of type 2 diabetes mellitus . Materials and Methods: The study was performed in 2 cohorts of populations, 112 healthy volunteers and 118 type 2 diabetes mellitus patients. Genomic deoxyribonucleic acid (DNA was extracted from peripheral blood leucocytes by phenol-chloroform method and genotyping was performed by real-time polymerase chain reaction using TaqMan assay. Results: In South Indian population, the frequency of major A allele was 0.65 and the minor C allele was 0.35. AA and CC are the homozygous genotypes with frequency of 0.39 and 0.09 respectively. The frequency of heterozygous genotype AC (0.52 was found to be higher than the homozygotes. There was no significant difference in the frequency distribution in the diabetic population, which implies that this variant does not have any causative role in the disease etiology. The frequency distributions were found to be significantly different from the distributions in other ethnic populations such as Caucasians, Chinese, Japanese and Africans. But there was no significant difference when compared with the Gujarati Indians of Houston. Conclusion: The frequency distribution of this novel variant in South Indian population forms a framework for further gene disease association studies to establish the association of this variant with metformin response. Our study could not find any association of this variant with respect to the disease etiology.

  17. Proteomic Characterization of Cerebrospinal Fluid from Ataxia-Telangiectasia (A-T Patients Using a LC/MS-Based Label-Free Protein Quantification Technology

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    Monika Dzieciatkowska

    2011-01-01

    Full Text Available Cerebrospinal fluid (CSF has been used for biomarker discovery of neurodegenerative diseases in humans since biological changes in the brain can be seen in this biofluid. Inactivation of A-T-mutated protein (ATM, a multifunctional protein kinase, is responsible for A-T, yet biochemical studies have not succeeded in conclusively identifying the molecular mechanism(s underlying the neurodegeneration seen in A-T patients or the proteins that can be used as biomarkers for neurologic assessment of A-T or as potential therapeutic targets. In this study, we applied a high-throughput LC/MS-based label-free protein quantification technology to quantitatively characterize the proteins in CSF samples in order to identify differentially expressed proteins that can serve as potential biomarker candidates for A-T. Among 204 identified CSF proteins with high peptide-identification confidence, thirteen showed significant protein expression changes. Bioinformatic analysis revealed that these 13 proteins are either involved in neurodegenerative disorders or cancer. Future molecular and functional characterization of these proteins would provide more insights into the potential therapeutic targets for the treatment of A-T and the biomarkers that can be used to monitor or predict A-T disease progression. Clinical validation studies are required before any of these proteins can be developed into clinically useful biomarkers.

  18. Chronic arsenic exposure increases TGFalpha concentration in bladder urothelial cells of Mexican populations environmentally exposed to inorganic arsenic☆

    Science.gov (United States)

    Valenzuela, Olga L.; Germolec, Dori R.; Borja-Aburto, Víctor H.; Contreras-Ruiz, José; García-Vargas, Gonzalo G.; Del Razo, Luz M.

    2009-01-01

    Inorganic arsenic (iAs) is a well-established carcinogen and human exposure has been associated with a variety of cancers including those of skin, lung, and bladder. High expression of transforming growth factor alpha (TGF-α) has associated with local relapses in early stages of urinary bladder cancer. iAs exposures are at least in part determined by the rate of formation and composition of iAs metabolites (MAsIII, MAsV, DMAsIII, DMAsV). This study examines the relationship between TGF-α concentration in exfoliated bladder urothelial cells (BUC) separated from urine and urinary arsenic species in 72 resident women (18-51 years old) from areas exposed to different concentrations of iAs in drinking water (2-378 ppb) in central Mexico. Urinary arsenic species, including trivalent methylated metabolites were measured by hydride generation atomic absorption spectrometry method. The concentration of TGF-α in BUC was measured using an ELISA assay. Results show a statistically significant positive correlation between TGF-α concentration in BUC and each of the six arsenic species present in urine. The multivariate linear regression analyses show that the increment of TGF-α levels in BUC was importantly associated with the presence of arsenic species after adjusting by age, and presence of urinary infection. People from areas with high arsenic exposure had a significantly higher TGF-α concentration in BUC than people from areas of low arsenic exposure (128.8 vs. 64.4 pg/mg protein; p<0.05). Notably, exfoliated cells isolated from individuals with skin lesions contained significantly greater amount of TGF-α than cells from individuals without skin lesions: 157.7 vs. 64.9 pg/mg protein (p=0.003). These results suggest that TGF-α in exfoliated BUC may serve as a susceptibility marker of adverse health effects on epithelial tissue in arsenic-endemic areas. PMID:17267001

  19. Differentiated THP-1 Cells Exposed to Pathogenic and Nonpathogenic Borrelia Species Demonstrate Minimal Differences in Production of Four Inflammatory Cytokines.

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    Stokes, John V; Moraru, Gail M; McIntosh, Chelsea; Kummari, Evangel; Rausch, Keiko; Varela-Stokes, Andrea S

    2016-11-01

    Tick-borne borreliae include Lyme disease and relapsing fever agents, and they are transmitted primarily by ixodid (hard) and argasid (soft) tick vectors, respectively. Tick-host interactions during feeding are complex, with host immune responses influenced by biological differences in tick feeding and individual differences within and between host species. One of the first encounters for spirochetes entering vertebrate host skin is with local antigen-presenting cells, regardless of whether the tick-associated Borrelia sp. is pathogenic. In this study, we performed a basic comparison of cytokine responses in THP-1-derived macrophages after exposure to selected borreliae, including a nonpathogen. By using THP-1 cells, differentiated to macrophages, we eliminated variations in host response and reduced the system to an in vitro model to evaluate the extent to which the Borrelia spp. influence cytokine production. Differentiated THP-1 cells were exposed to four Borrelia spp., Borrelia hermsii (DAH), Borrelia burgdorferi (B31), B. burgdorferi (NC-2), or Borrelia lonestari (LS-1), or lipopolysaccharides (LPS) (activated) or media (no treatment) controls. Intracellular and secreted interferon (IFN)-γ, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α were measured using flow cytometric and Luminex-based assays, respectively, at 6, 24, and 48 h postexposure time points. Using a general linear model ANOVA for each cytokine, treatment (all Borrelia spp. and LPS compared to no treatment) had a significant effect on secreted TNF-α only. Time point had a significant effect on intracellular IFN-γ, TNF-α and IL-6. However, we did not see significant differences in selected cytokines among Borrelia spp. Thus, in this model, we were unable to distinguish pathogenic from nonpathogenic borreliae using the limited array of selected cytokines. While unique immune profiles may be detectable in an in vitro model and may reveal predictors for pathogenicity in borreliae

  20. NPRL2 sensitizes human non-small cell lung cancer (NSCLC cells to cisplatin treatment by regulating key components in the DNA repair pathway.

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    Gitanjali Jayachandran

    2010-08-01

    Full Text Available NPRL2, one of the tumor suppressor genes residing in a 120-kb homozygous deletion region of human chromosome band 3p21.3, has a high degree of amino acid sequence homology with the nitrogen permease regulator 2 (NPR2 yeast gene, and mutations of NPRL2 in yeast cells are associated with resistance to cisplatin-mediated cell killing. Previously, we showed that restoration of NPRL2 in NPRL2-negative and cisplatin-resistant cells resensitize lung cancer cells to cisplatin treatment in vitro and in vivo. In this study, we show that sensitization of non-small cell lung cancer (NSCLC cells to cisplatin by NPRL2 is accomplished through the regulation of key components in the DNA-damage checkpoint pathway. NPRL2 can phosphorylate ataxia telangiectasia mutated (ATM kinase activated by cisplatin and promote downstream gamma-H2AX formation in vitro and in vivo, which occurs during apoptosis concurrently with the initial appearance of high-molecular-weight DNA fragments. Moreover, this combination treatment results in higher Chk1 and Chk2 kinase activity than does treatment with cisplatin alone and can activate Chk2 in pleural metastases tumor xenograft in mice. Activated Chk1 and Chk2 increase the expression of cell cycle checkpoint proteins, including Cdc25A and Cdc25C, leading to higher levels of G2/M arrest in tumor cells treated with NPRL2 and cisplatin than in tumor cells treated with cisplatin only. Our results therefore suggest that ectopic expression of NPRL2 activates the DNA damage checkpoint pathway in cisplatin-resistant and NPRL2-negative cells; hence, the combination of NPRL2 and cisplatin can resensitize cisplatin nonresponders to cisplatin treatment through the activation of the DNA damage checkpoint pathway, leading to cell arrest in the G2/M phase and induction of apoptosis. The direct implication of this study is that combination treatment with NPRL2 and cisplatin may overcome cisplatin resistance and enhance therapeutic efficacy.

  1. NPRL2 Sensitizes Human Non-Small Cell Lung Cancer (NSCLC) Cells to Cisplatin Treatment by Regulating Key Components in the DNA Repair Pathway

    Science.gov (United States)

    Wang, Bingbing; Roth, Jack A.; Ji, Lin

    2010-01-01

    NPRL2, one of the tumor suppressor genes residing in a 120-kb homozygous deletion region of human chromosome band 3p21.3, has a high degree of amino acid sequence homology with the nitrogen permease regulator 2 (NPR2) yeast gene, and mutations of NPRL2 in yeast cells are associated with resistance to cisplatin-mediated cell killing. Previously, we showed that restoration of NPRL2 in NPRL2-negative and cisplatin-resistant cells resensitize lung cancer cells to cisplatin treatment in vitro and in vivo. In this study, we show that sensitization of non-small cell lung cancer (NSCLC) cells to cisplatin by NPRL2 is accomplished through the regulation of key components in the DNA-damage checkpoint pathway. NPRL2 can phosphorylate ataxia telangiectasia mutated (ATM) kinase activated by cisplatin and promote downstream γ-H2AX formation in vitro and in vivo, which occurs during apoptosis concurrently with the initial appearance of high-molecular-weight DNA fragments. Moreover, this combination treatment results in higher Chk1 and Chk2 kinase activity than does treatment with cisplatin alone and can activate Chk2 in pleural metastases tumor xenograft in mice. Activated Chk1 and Chk2 increase the expression of cell cycle checkpoint proteins, including Cdc25A and Cdc25C, leading to higher levels of G2/M arrest in tumor cells treated with NPRL2 and cisplatin than in tumor cells treated with cisplatin only. Our results therefore suggest that ectopic expression of NPRL2 activates the DNA damage checkpoint pathway in cisplatin-resistant and NPRL2-negative cells; hence, the combination of NPRL2 and cisplatin can resensitize cisplatin nonresponders to cisplatin treatment through the activation of the DNA damage checkpoint pathway, leading to cell arrest in the G2/M phase and induction of apoptosis. The direct implication of this study is that combination treatment with NPRL2 and cisplatin may overcome cisplatin resistance and enhance therapeutic efficacy. PMID:20700484

  2. Methods for exposing multiple cultures of endothelial cells to different fluid shear stresses and to cytokines, for subsequent analysis of inflammatory function.

    Science.gov (United States)

    Sheikh, Sajila; Gale, Zoe; Rainger, G Ed; Nas